New cultivation strategies bring more microbial plankton species into the laboratory (original) (raw)
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Frontiers in Microbiology, 2014
Some microalgae in nature live in symbiosis with microorganisms that can enhance or inhibit growth, thus influencing the dynamics of phytoplankton blooms. In spite of the great ecological importance of these interactions, very few defined laboratory systems are available to study them in detail. Here we present a co-cultivation system consisting of the toxic phototrophic dinoflagellate Prorocentrum minimum and the photoheterotrophic alphaproteobacterium Dinoroseobacter shibae. In a mineral medium lacking a carbon source, vitamins for the bacterium and the essential vitamin B 12 for the dinoflagellate, growth dynamics reproducibly went from a mutualistic phase, where both algae and bacteria grow, to a pathogenic phase, where the algae are killed by the bacteria. The data show a "Jekyll and Hyde" lifestyle that had been proposed but not previously demonstrated. We used RNAseq and microarray analysis to determine which genes of D. shibae are transcribed and differentially expressed in a light dependent way at an early time-point of the co-culture when the bacterium grows very slowly. Enrichment of bacterial mRNA for transcriptome analysis was optimized, but none of the available methods proved capable of removing dinoflagellate ribosomal RNA completely. RNAseq showed that a phasin encoding gene (phaP 1) which is part of the polyhydroxyalkanoate (PHA) metabolism operon represented approximately 10% of all transcripts. Five genes for aerobic anoxygenic photosynthesis were down-regulated in the light, indicating that the photosynthesis apparatus was functional. A betaine-choline-carnitine-transporter (BCCT) that may be used for dimethylsulfoniopropionate (DMSP) uptake was the highest up-regulated gene in the light. The data suggest that at this early mutualistic phase of the symbiosis, PHA degradation might be the main carbon and energy source of D. shibae, supplemented in the light by degradation of DMSP and aerobic anoxygenic photosynthesis.
FEMS Microbiology Ecology, 2011
The prokaryotic activity, diversity and culturability of diffusion-controlled Aarhus Bay sediments, including the sulphate-methane transition zone (SMTZ), were determined using a combination of geochemical, molecular (16S rRNA and mcrA genes) and cultivation techniques. The SMTZ had elevated sulphate reduction and anaerobic oxidation of methane, and enhanced cell numbers, but no active methanogenesis. The prokaryotic population was similar to that in other SMTZs, with Deltaproteobacteria, Gammaproteobacteria, JS1, Planctomycetes, Chloroflexi, ANME-1, MBG-D and MCG. Many of these groups were maintained in a heterotrophic (10 mM glucose, acetate), sediment slurry with periodic low sulphate and acetate additions ($2 mM). Other prokaryotes were also enriched including methanogens, Firmicutes, Bacteroidetes, Synergistetes and TM6. This slurry was then inoculated into a matrix of substrate and sulphate concentrations for further selective enrichment. The results demonstrated that important SMTZ bacteria can be maintained in a long-term, anaerobic culture under specific conditions. For example, JS1 grew in a mixed culture with acetate or acetate/ glucose plus sulphate. Chloroflexi occurred in a mixed culture, including in the presence of acetate, which had previously not been shown to be a Chloroflexi subphylum I substrate, and was more dominant in a medium with seawater salt concentrations. In contrast, archaeal diversity was reduced and limited to the orders Methanosarcinales and Methanomicrobiales. These results provide information about the physiology of a range of SMTZ prokaryotes and shows that many can be maintained and enriched under heterotrophic conditions, including those with few or no cultivated representatives.
Physiology, Ecology, Phylogeny, and Genomics of Microorganisms Capable of Syntrophic Metabolism
Annals of the New York Academy of Sciences, 2008
Syntrophic metabolism is diverse in two respects: phylogenetically with microorganisms capable of syntrophic metabolism found in the Deltaproteobacteria and in the low G+C gram-positive bacteria, and metabolically given the wide variety of compounds that can be syntrophically metabolized. The latter includes saturated fatty acids, unsaturated fatty acids, alcohols, and hydrocarbons. Besides residing in freshwater and marine anoxic sediments and soils, microbes capable of syntrophic metabolism also have been observed in more extreme habitats, including acidic soils, alkaline soils, thermal springs, and permanently cold soils, demonstrating that syntrophy is a widely distributed metabolic process in nature. Recent ecological and physiological studies show that syntrophy plays a far larger role in carbon cycling than was previously thought. The availability of the first complete genome sequences for four model microorganisms capable of syntrophic metabolism provides the genetic framework to begin dissecting the biochemistry of the marginal energy economies and interspecies interactions that are characteristic of the syntrophic lifestyle.
A mini-review of microbial consortia: Their roles in aquatic production and biogeochemical cycling
Microbial Ecology, 1996
Molecular oxygen (02) is a potent inhibitor of key microbial processes, including photosynthesis, N2 fixation, denitrification, sulfate reduction, methanogenesis, iron, and metal reduction reactions. Prokaryote survival and proliferation in aquatic environments is often controlled by the ability to tolerate exposure to oxic conditions. Many prokaryotes do not have subcellular organelles for isolating O2-producing from O2-consuming processes and have developed consortial associations with other prokaryotes and eukaryotes that alleviate metabolic constraints of high 02. Nutrient transformations often rely on appropriate cellular and microenvironmental, or microzonal, redox conditions. The spatial and temporal requirements for microenvironmental overlap among microbial groups involved in nutrient transformations necessitates close proximity and diffusional exchange with other biogeochemically distinct, yet complementary, microbial groups. Microbial consortia exist at different levels of community and metabolic complexity, as shown for detrital, microbial mat, biofilm, and planktonic microalgal-bacterial assemblages. To assess the macroscale impacts of consortial interactions, studies should focus on the range of relevant temporal (minutes to hours) and spatial (microns to centimeters) scales controlling microbial production, nutrient exchange, and cycling. In this review, we discuss the utility and application of techniques suitable for determining microscale consortial activity, production, community composition, and interactions in the context of larger scale aquatic ecosystem structure and function.
PeerJ, 2015
We have previously observed that methane supplied to lake sediment microbial communities as a substrate not only causes a response by bona fide methanotrophic bacteria, but also by non-methane-oxidizing bacteria, especially by members of the family Methylophilaceae. This result suggested that methane oxidation in this environment likely involves communities composed of different functional guilds, rather than a single type of microbe. To obtain further support for this concept and to obtain further insights into the factors that may define such partnerships, we carried out microcosm incubations with sediment samples from Lake Washington at five different oxygen tensions, while methane was supplied at the same concentration in each. Community composition was determined through 16S rRNA gene amplicon sequencing after 10 and 16 weeks of incubation. We demonstrate that, in support of our prior observations, the methane-consuming communities were represented by two major types: the methanotrophs of the family Methylococcaceae and by non-methanotrophic methylotrophs of the family Methylophilaceae. However, different species persisted under different oxygen tensions. At high initial oxygen tensions (150 to 225 µM) the major players were, respectively, species of the genera Methylosarcina and Methylophilus, while at low initial oxygen tensions (15 to 75 µM) the major players were Methylobacter and Methylotenera. These data suggest that oxygen availability is at least one major factor determining specific partnerships in methane oxidation. The data also suggest that speciation within Methylococcaceae and Methylophilaceae may be driven by niche adaptation tailored toward specific placements within the oxygen gradient. How to cite this article Hernandez et al. (2015), Oxygen availability is a major factor in determining the composition of microbial communities involved in methane oxidation. PeerJ 3:e801; -Tringe S, Grigoriev I, Markowitz V, Rigoutsos I, Richardson PM, Lidstrom ME, Chistoserdova L. 2008. High resolution metagenomics targets major functional types in complex microbial communities. Nature Biotechnology 26:1029-1034 DOI 10.1038/nbt.1488. Kalyuzhnaya MG, Stolyar SM, Auman AJ, Lara JC, Lidstrom ME, Chistoserdova L. 2005. Methylosarcina lacus sp. nov., a methanotroph from Lake Washington, Seattle, USA, and emended description of the genus Methylosarcina. International Journal of Systematic and Evolutionary Microbiology 55:2345-2350 DOI 10.1099/ijs.0.63405-0. Kits KD, Klotz MG, Stein LY. 2015. Methane oxidation coupled to nitrate reduction under hypoxia by the Gammaproteobacterium Methylomonas denitrificans, sp. nov. Type Strain FJG1. Environmental Microbiology . Mustakhimov I, Kalyuzhnaya MG, Lidstrom MW, Chistoserdova L. 2013. Insights into denitrification in Methylotenera mobilis from denitrification pathway and methanol metabolism mutants. . Tavormina PL, Orphan VJ, Kalyuzhnaya MG, Jetten MS, Klotz MG. 2011. A novel family of functional operons encoding methane/ammonia monooxygenase-related proteins in gammaproteobacterial methanotrophs. Environmental Microbiology Reports 3:91-100
Characterization of a marine gammaproteobacterium capable of aerobic anoxygenic photosynthesis
Proceedings of the National Academy of Sciences, 2007
Members of the gammaproteobacterial clade NOR5/OM60 regularly form an abundant part, up to 11%, of the bacterioplankton community in coastal systems during the summer months. Here, we report the nearly complete genome sequence of one cultured representative, Congregibacter litoralis strain KT71, isolated from North Sea surface water. Unexpectedly, a complete photosynthesis superoperon, including genes for accessory pigments, was discovered. It has a high sequence similarity to BAC clones from Bay [Beja O, Suzuki MT, Heidelberg JF, Nelson WC, Preston CM, et al. (2002) Nature 415:630 -633], which also share a nearly identical gene arrangement. Although cultures of KT71 show no obvious pigmentation, bacteriochlorophyll a and spirilloxanthin-like carotenoids could be detected by HPLC analysis in cell extracts. The presence of two potential BLUF (blue light using flavin adenine dinucleotide sensors), one of which was found adjacent to the photosynthesis operon in the genome, indicates a light-and redox-dependent regulation of gene expression. Like other aerobic anoxygenic phototrophs (AAnPs), KT71 is able to grow neither anaerobically nor photoautotrophically. Cultivation experiments and genomic evidence show that KT71 needs organic substrates like carboxylic acids, oligopeptides, or fatty acids for growth. The strain grows optimally under microaerobic conditions and actively places itself in a zone of Ϸ10% oxygen saturation. The genome analysis of C. litoralis strain KT71 identifies the gammaproteobacterial marine AAnPs, postulated based on BAC sequences, as members of the NOR5/OM60 clade. KT71 enables future experiments investigating the importance of this group of gammaproteobacterial AAnPs in coastal environments. marine bacteria ͉ NOR5/OM60 clade ͉ whole genome analysis ͉ Congregibacter litoralis ͉ bacteriochlorophyll a
Aquatic Microbial Ecology 38:203
2005
The effect of different phosphorus loads (L P) on the phosphorus (P) and carbon (C) content (biomass) of algae and bacteria was assessed in continuous culture. We tested if a mixed freshwater microbial assemblage co-cultured with a phytoflagellate (Cryptomonas phaseolus) would comply with the 'phytoplankton-bacteria paradox' (sensu Bratbak & Thingstad 1985). This hypothesis states that the ratio of bacterial to algal abundance changes to the benefit of bacteria with decreasing L P. However, the phenomenon was originally investigated by simultaneously altering L P and microbial growth rates, and it is unclear to which extent it can be assigned to either parameter. Therefore, we set up 3 chemostat systems in triplicate at equal dilution rates, but with daily L P of 21, 41 or 62 µg l-1 d-1 (corresponding to 50, 100 and 150 µg P l-1). Higher L P led to a 5-fold increase in total algal abundance and biomass but to less than a doubling of these parameters in the bacterial assemblage. Total biomass ratios of bacteria to algae changed from 0.18 to 0.06 with increasing L P , while the bacteria-algae total phosphorus ratios decreased from 0.80 to 0.17. The cellular C:P ratio of algae remained similar at all P concentrations, whereas the molar C:P ratios of bacterial cells significantly increased at higher L P (from 44 to 73). An enrichment experiment with the 50 µg P l-1 treatment demonstrated that bacteria at the lowest L P were co-limited by P and C, and that increased P stimulated mainly the algal fraction. The phytoplankton-bacteria paradox at the level of a mixed microbial assemblage is thus characterised by the following aspects: (1) bacteria profit from their high affinity to P and are better competitors at lower L P ; (2) although algae compete with bacteria for P, P-limited algae release extracellular C that stimulates growth of their bacterial competitors; (3) when bacteria depend on algae as their sole source of organic C, this provides a feedback mechanism by which algae limit the abundance of their competitors at higher L P ; (4) large oscillations in the bacteria-algae ratios at the lowest L P point to a greater instability of this interaction with stronger P competition. However, bacteria were not able to outcompete C. phaseolus, as algae were their only C source.
Ophelia, 1993
Anaerobic ciliates with endosymbiotic methanogenic bacteria are sometimes responsible for a substantial fraction (15-90%) of the production of CHi in anaerobic detrital sediments and in the anoxic water column. In sandy sediments anaerobic protozoa playa relatively smaller role (typically < 2%) due to their low numbers and limited vertical distribution and since there is a relatively large production of methane by free-living methanogens associated with detrital particles even in the upper, sulphate-rich layers of the sediment. Under all circumstances protozoa playa small role for the terminal mineralisation in anaerobic biota since phagotrophs represent a second trophic level in an ecosystem with low growth efficiencies. In sulphate-poor anoxic habitats endosymbiotic methanogenesis wiu therefore always playa relatively small quantitative role.
Mesozooplankton provide oxic and anoxic microhabitats for associated bacteria, whose carbon substrate usage activities complement those of the ambient bacteria. The metabolic profiles of bacterial communities associated with the calanoid copepod Acartia tonsa under aerobic and anaerobic conditions were examined in comparison with phytoplankton-associated bacteria. Carbon substrate usage by phytoplankton-associated bacteria was significantly different than that of copepod-associated bacteria in both aerobic and anaerobic conditions. Substrate utilization by copepod-associated bacteria was more dependent upon oxygen condition than whether the bacteria were located on the copepod exoskeleton or within the gut. Results suggest that gut bacteria were responsible for a large portion of anaerobic substrate usage by copepod-associated bacteria. The metabolic profiles of bacteria associated with six common zooplankton groups and free-living bacteria collected in July 2012 from the York River estuary, Virginia, (37°14′50.36″N, 76°29′58.03W) were also compared, and there were significant differences in their substrate utilization patterns between aerobic and anaerobic incubations, and among the different zooplankton groups. Through trophic interactions, phytoplankton-associated or free-living bacteria may be introduced to the anoxic zooplankton gut and its associated bacterial community. Inclusion of these anaerobic microenvironments and their microbial inhabitants increased the total number of substrates used by 57 % over what was used by aerobic phytoplankton-associated bacteria alone, and by 50 % over what was used by aerobic free-living bacteria in the York River. Therefore, the presence of zooplankton-associated microhabitats and their bacteria expanded the functionality of aquatic microbial communities and led to a more comprehensive substrate usage.
Minireview A Great Leap forward in Microbial Ecology
Ribosomal RNA (rRNA) sequence-based molecular techniques emerged in the late 1980s, which completely changed our general view of microbial life. Coincidentally, the Japanese Society of Microbial Ecology (JSME) was founded, and its official journal " Microbes and Environments (M&E) " was launched, in 1985. Thus, the past 25 years have been an exciting and fruitful period for M&E readers and microbiologists as demonstrated by the numerous excellent papers published in M&E. In this minireview, recent progress made in microbial ecology and related fields is summarized, with a special emphasis on 8 landmark areas; the cultivation of uncultured microbes, in situ methods for the assessment of microorganisms and their activities, biofilms, plant microbiology, chemolithotrophic bacteria in early volcanic environments, symbionts of animals and their ecology, wastewater treatment microbiology, and the biodegradation of hazardous organic compounds.