Bram Vekeman

Safeguarding bacterial resources promotes biotechnological innovation

Environmental research delivers valuable bacterial resources for biotechnology. We believe that systematic long-term preservation of bacteria will promote future biotechnological innovations, by safeguarding the accessibility of bacteria already recognized to have interesting features and providing a “pool” of bacterial resources for novel applied research. To this end, we want to advocate the incorporation of preservation tests in environmental or applied microbiological research. This paper introduces non-specialists to different preservation methods for bacteria. Several parameters that influence long-term storage of bacterial resources are explained and practical tips and guidelines are formulated. Also, the vital role of public culture collections is highlighted and the state-of-the-art of preservation of non-pure cultures is described.

A generally applicable cryopreservation method for nitrite-oxidizing bacteria

Nitrite-oxidizing bacteria are key members of the global nitrogen cycle but their study is hampered by their limited availability in culture, mostly due to laborious cultivation procedures and the lack of stable preservation methods. In this study, it was demonstrated that long-term cryopreservation of nitrite-oxidizing bacteria assigned to the genera Nitrobacter, Nitrospina, Nitrococcus, Nitrotoga and Nitrospira was possible using a simple and rapid protocol. Their survival was tested with different cryoprotecting agents, DMSO and Hatefi, and in various carbon-rich preservation media, ten-fold diluted TSB, and ten-fold diluted TSB supplemented with 1% trehalose, and 1% sucrose. Optimal preservation conditions were strain-dependent and marine strains appeared to be more sensitive to freezing than non-marine strains. Nevertheless, a general cryopreservation protocol using 10% dimethyl sulfoxide with or without ten-fold diluted trypticase soy broth as a preservation medium allowed successful preservation of all tested strains.

Draft Genome Sequences of Eight Obligate Methane Oxidizers Occupying Distinct Niches Based on Their Nitrogen Metabolism.

ABSTRACT The genome sequences of Methylomonas methanica (NCIMB 11130T, R-45363, and R-45371), Methylomonas koyamae (R-45378, R-45383, and R-49807), Methylomonas lenta (R-45370), and Methylosinus sp. (R-45379) were obtained. These aerobic methanotrophs were isolated from terrestrial ecosystems, and their distinct phenotypes related to nitrogen assimilation and dissimilation were previously reported.

New Methyloceanibacter diversity from North Sea sediments includes methanotroph containing solely the soluble methane monooxygenase

Marine methylotrophs play a key role in the global carbon cycle by metabolizing reduced one-carbon compounds that are found in high concentrations in marine environments. Genome, physiology and diversity studies have been greatly facilitated by the numerous model organisms brought into culture. However, the availability of marine representatives remains poor. Here, we report the isolation of four novel species from North Sea sediment enrichments closely related to the Alphaproteobacterium Methyloceanibacter caenitepidi. Each of the newly isolated Methyloceanibacter species exhibited a clear genome sequence divergence which was reflected in physiological differences. Notably one strain R-67174 was capable of oxidizing methane as sole source of carbon and energy using solely a soluble methane monooxygenase and represents the first marine Alphaproteobacterial methanotroph brought into culture. Differences in maximum cell density of >1.5 orders of magnitude were observed. Furthermore, three strains were capable of producing nitrous oxide from nitrate. Together, these findings highlight the metabolic and physiologic variability within closely related Methyloceanibacter species and provide a new understanding of the physiological basis of marine methylotrophy.

Improved enrichment culture technique for methane-oxidizing bacteria from marine ecosystems: the effect of adhesion material and gas composition

Cultivation of microbial representatives of specific functional guilds from environmental samples depends largely on the suitability of the applied growth conditions. Especially the cultivation of marine methanotrophs has received little attention, resulting in only a limited number of ex situ cultures available. In this study we investigated the effect of adhesion material and headspace composition on the methane oxidation activity in methanotrophic enrichments obtained from marine sediment. Addition of sterilized natural sediment or alternatively the addition of acid-washed silicon dioxide significantly increased methane oxidation. This positive effect was attributed to bacterial adhesion on the particles via extracellular compounds, with a minimum amount of particles required for effect. As a result, the particles were immobilized, thus creating a stratified environment in which a limited diffusive gas gradients could build up and various microniches were formed. Such diffusive gas gradient might necessitate high headspace concentrations of CH4 and CO2 for sufficient concentrations to reach the methane-oxidizing bacteria in the enrichment culture technique. Therefore, high concentrations of methane and carbon dioxide, in addition to the addition of adhesion material, were tested and indeed further stimulated methane oxidation. Use of adhesion material in combination with high concentrations of methane and carbon dioxide might thus facilitate the cultivation and subsequent enrichment of environmentally important members of this functional guild. The exact mechanism of the observed positive effects on methane oxidation and the differential effect on methanotrophic diversity still needs to be explored.