title: A novel method for exploring elemental composition of microbial communities: laser ablation-inductively coupled plasma-mass spectrometry of intact bacterial colonies.
abstract: Bacterial colonies are spatially complex structures whose physiology is profoundly dependent on interactions between cells and with the underlying semi-solid substratum. Here, we use bacterial colonies as a model of a microbial community to evaluate the potential of laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) to delineate elemental distributions within colonies with minimal pre-treatment. To reduce water content of the colony and limit undesirable absorption of laser energy, we compared methods of preparing 24h-old colonies of Escherichia coli TG1 on agar for laser ablation. Colonies on excised agar segments dried on chromatography paper were superior to colonies dried in a dessicator or by prolonged incubation, with respect to signal magnitude, signal:noise ratio and background signal. Having optimised laser scan speed (10 microm s(-1)) and laser beam diameter (100 microm), further improvements were achieved by growing colonies on nylon membranes over agar, which were then transferred to the ablation chamber without further treatment. Repeated line rasters across individual membrane-supported colonies yielded three-dimensional elemental maps of colonies, revealing a convex morphology consistent with visual inspection. By normalising isotope counts for P, Mn, Zn, Fe and Ca against Mg, the most abundant cellular divalent cation, we sought elemental heterogeneity within the colony. The normalised concentration of Mn in the perimeter was higher than in the colony interior, whereas the converse was true for Ca. LA-ICP-MS is a novel and powerful method for probing elemental composition and organisation within microbial communities and should find numerous applications in, for example, biofilm studies.
authors.affiliation: Department of Molecular Biology and Biotechnology, Firth Court, Western Bank, The University of Sheffield, Sheffield S10 TN, UK.
authors.forename: Joe
authors.initials: J
authors.lastname: Latimer
authors.forename: Sarah L
authors.initials: SL
authors.lastname: Stokes
authors.forename: Alison I
authors.initials: AI
authors.lastname: Graham
authors.forename: Josephine
authors.initials: J
authors.lastname: Bunch
authors.forename: Rachel J
authors.initials: RJ
authors.lastname: Jackson
authors.forename: Cameron W
authors.initials: CW
authors.lastname: McLeod
authors.forename: Robert K
authors.initials: RK
authors.lastname: Poole
mesh_terms: D001419:Bacteria / Q000737:chemistry*; D018407:Bacterial Physiological Phenomena; D018441:Biofilms; D004926:Escherichia coli / Q000254:growth & development; D007834:Lasers; D013058:Mass Spectrometry / Q000379:methods*; D008567:Membranes, Artificial; D008851:Micropore Filters; D008954:Models, Biological; D009757:Nylons
chemical_list: D008567:Membranes, Artificial; D009757:Nylons