Breast milk-derived human milk oligosaccharides promote Bifidobacterium interactions within a single ecosystem

Diet-microbe interactions play an important role in modulating the early-life microbiota, with Bifidobacterium strains and species dominating the gut of breast-fed infants. Here, we sought to explore how infant diet drives distinct bifidobacterial community composition and dynamics within individual infant ecosystems. Genomic characterisation of 19 strains isolated from breast-fed infants revealed a diverse genomic architecture enriched in carbohydrate metabolism genes, which was distinct to each strain, but collectively formed a pangenome across infants. Presence of gene clusters implicated in digestion of human milk oligosaccharides (HMOs) varied between species, with growth studies indicating that within single infants there were differences in the ability to utilise 2'FL and LNnT HMOs between strains. Cross-feeding experiments were performed with HMO degraders and non-HMO users (using spent or 'conditioned' media and direct co-culture). Further H-NMR analysis identified fucose, galactose, acetate, and N-acetylglucosamine as key by-products of HMO metabolism; as demonstrated by modest growth of non-HMO users on spend media from HMO metabolism. These experiments indicate how HMO metabolism permits the sharing of resources to maximise nutrient consumption from the diet and highlights the cooperative nature of bifidobacterial strains and their role as 'foundation' species in the infant ecosystem. The intra- and inter-infant bifidobacterial community behaviour may contribute to the diversity and dominance of Bifidobacterium in early life and suggests avenues for future development of new diet and microbiota-based therapies to promote infant health

SIGNALING STUDIES IN THE EMERGING KIWIFRUIT PATHOGEN Pseudomonas syringae pv. actinidiae

In the past two decades emerging and re-emerging plant pathogens have caused new threats to the production of several economically important crops, one among them is P. syringae pv. actinidiae (PSA) which causes canker or leaf spot on kiwifruit plants. PSA enters plant through wounds and remains dormant in cortex tissue of the branches, and spreads in the tissue to cause severe symptoms from winter to early spring. The disease can be visualized by brown discoloration of buds, dark brown angular spots surrounded by yellow haloes on leaves, cankers with white to reddish (oxydation) exudate on twigs and trunks, fruit collapse, wilting and eventually plant mortality. Current control methods have their own significance in disease control, however there is considerable lack of clear understanding of PSA pathogenicity. Virulence of plant pathogens often relies on the synchronized/coordinated expression of pathogenicity factors via quorum sensing (QS). Therefore, investigations on QS in PSA may lead to develop novel disease control strategies and reliable methods to curb the disease. It is currently unknown whether PSA produces a QS signal molecule thus the aim of this thesis is to investigate whether PSA possesses a QS system. As genome mining did not reveal the presence of any currently known QS system, this study initially by metabolomics was aimed at identifying potentially low molecular weight secondary metabolite QS molecules produced by PSA. Azelaic acid was discovered to be produced by PSA, this is the first report of azelaic acid production by bacteria. The characterization and possible role of azelaic acid in QS is presented. Since azelaic acid is ubiquitous in nature, in addition to determining its biological role, the catabolism of azelaic acid in bacteria using the efficient degrader Pseudomonas nitroreducens DSM 9128 was also studied

Identification of loci of pseudomonas syringae pv. actinidiae involved in lipolytic activity and their role in colonization of kiwifruit leaves

Bacterial canker disease caused by Pseudomonas syringae pv. actinidiae, an emerging pathogen of kiwifruit plants, has recently brought about major economic losses worldwide. Genetic studies on virulence functions of P. syringae pv. actinidiae have not yet been reported and there is little experimental data regarding bacterial genes involved in pathogenesis. In this study, we performed a genetic screen in order to identify transposon mutants altered in the lipolytic activity because it is known that mechanisms of regulation, production, and secretion of enzymes often play crucial roles in virulence of plant pathogens. We aimed to identify the set of secretion and global regulatory loci that control lipolytic activity and also play important roles in in planta fitness. Our screen for altered lipolytic activity phenotype identified a total of 58 Tn5 transposon mutants. Mapping all these Tn5 mutants revealed that the transposons were inserted in genes that play roles in cell division, chemotaxis, metabolism, movement, recombination, regulation, signal transduction, and transport as well as a few unknown functions. Several of these identified P. syringae pv. actinidiae Tn5 mutants, notably the functions affected in phosphomannomutase AlgC, lipid A biosynthesis acyltransferase, glutamate–cysteine ligase, and the type IV pilus protein PilI, were also found affected in in planta survival and/or growth in kiwifruit plants. The results of the genetic screen and identification of novel loci involved in in planta fitness of P. syringae pv. actinidiae are presented and discussed

The spent culture supernatant of Pseudomonas syringae contains azelaic acid

Abstract Background Pseudomonas syringae pv. actinidiae (PSA) is an emerging kiwifruit bacterial pathogen which since 2008 has caused considerable losses. No quorum sensing (QS) signaling molecule has yet been reported from PSA and the aim of this study was to identify possible intercellular signals produced by PSA. Results A secreted metabolome analysis resulted in the identification of 83 putative compounds, one of them was the nine carbon saturated dicarboxylic acid called azelaic acid. Azelaic acid, which is a nine-carbon (C9) saturated dicarboxylic acid, has been reported in plants as a mobile signal that primes systemic defenses. In addition, its structure,(which is associated with fatty acid biosynthesis) is similar to other known bacterial QS signals like the Diffusible Signal Facor (DSF). For these reason it could be acting as s signal molecule. Analytical and structural studies by NMR spectroscopy confirmed that in PSA spent supernatants azelaic acid was present. Quantification studies further revealed that 20 μg/L of were present and was also found in the spent supernatants of several other P. syringae pathovars. The RNAseq transcriptome study however did not determine whether azelaic acid could behave as a QS molecule. Conclusions This study reports of the possible natural biosynthesis of azelaic acid by bacteria. The production of azelaic acid by P. syringae pathovars can be associated with plant-bacteria signaling