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Bacterial lifestyle switch in response to algal metabolites

ABSTRACT: Unicellular algae, termed phytoplankton, greatly impact the marine environment by serving as the basis of marine food webs and by playing central roles in biogeochemical cycling of elements. The interactions between phytoplankton and heterotrophic bacteria affect the fitness of both partners. It is becoming increasingly understood that metabolic exchange determines the nature of such interactions, but the underlying molecular mechanisms remain underexplored. Here, we investigated the molecular and metabolic basis for the bacterial lifestyle switch, from coexistence to pathogenicity, in Sulfitobacter D7 during interactions with Emiliania huxleyi, a cosmopolitan bloom-forming phytoplankter. The interaction displays two distinct phases: first, there is a coexisting phase in which the alga grows exponentially and the bacterium grows as well. The interaction shifts to pathogenic when the virulence of Sulfitobacter D7 towards E. huxleyi is invoked upon exposure to high concentrations of algal dimethylsulfoniopropionate (DMSP), which occurs when the algae reach stationary growth or when DMSP is applied exogenously to algae in exponential growth. We aimed to unravel the response of Sulfitobacter D7 to the pathogenicity-inducing compound, DMSP, and to different algae-derived infochemicals that affect the lifestyle of the bacterium. We grew Sulfitobacter D7 in conditioned media (CM) derived from algal cultures at the different growth phases, exponential and stationary (Exp-CM and Stat-CM, respectively), in which DMSP concentration is low and high, respectively. This enabled us to separate between different phases of the interaction with E. huxleyi, i.e., Exp-CM representing the coexisting phase, and Stat-CM representing the pathogenic phase. An additional pathogenicity-inducing treatment was Exp-CM supplemented with 100 µM DMSP (herein Exp-CM+DMSP). This condition mimicked co-cultures to which we added DMSP exogenously and thus induced Sulfitobacter D7 pathogenicity, which lead to death of exponentially growing E. huxleyi. In order to identify bacterial genes that are specifically responsive to DMSP, and are not affected by other algae-derived factors, we grew Sulfitobacter D7 in defined minimal medium (MM), lacking algal metabolites, supplemented with 100 µM DMSP (herein MM+DMSP), and examined the transcriptional response. After 24 h of Sulfitobacter D7 growth in all 5 media, triplicates were taken for transcriptomic analysis. Altogether, this experimental design allowed to expand our understanding on the bacterial response to DMSP, algal infochemicals and which of these are essential for coexistence and pathogenicity.

ORGANISM(S): Sulfitobacter sp. D7

PROVIDER: GSE193203 | GEO | 2023/01/17

REPOSITORIES: GEO

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Project description:The fraction of dissolved dimethylsulfoniopropionate (DMSPd) converted by marine bacterioplankton into the climate-active gas dimethylsulfide (DMS) varies widely in the ocean, with the factors that determine this value still largely unknown. One current hypothesis is that the ratio of DMS formation:DMSP demethylation is determined by DMSP availability, with 'availability' in both an absolute sense (i.e., concentration in seawater) and in a relative sense (i.e., proportionally to other labile organic S compounds) being proposed as the critical factor. We investigated these models during an experimentally-induced phytoplankton bloom using an environmental microarray targeting DMSP-related gene expression in the Roseobacter group, a taxon of marine bacteria known to play an important role in the surface ocean sulfur cycle. The array consisted of 1,578 probes to 431 genes, including those previously linked to DMSP degradation as well as core genes common in sequenced Roseobacter genomes. The prevailing pattern of Roseobacter gene expression showed depletion of DMSP-related transcripts during the peak of the bloom, despite the fact that absolute concentrations and flux of DMSP-related compounds were increasing. A likely interpretation is that DMSPd was assimilated by Roseobacter populations in proportion to its relative abundance in the organic matter pool (the “relative sense” hypothesis), and that it is not taken up in preference to other sources of labile organic sulfur or carbon produced during the bloom. The relative investment of the Roseobacter community in DMSP demethylation did not predict the fractional conversion of DMSP to DMS, however, suggesting a complex regulatory process that may involve multiple fates of DMSPd. DMSP-related gene expression in the Roseobacter group was investigated using an environmental microarray. Coastal seawater from the Gulf of Mexico was collected and dispensed into 20-L microcosms. Two replicate cubitainers were amended with nutrients (N and P) to stimulate phytoplankton bloom, while two untreated cubitainers served as controls. The microcosms were incubated at 27ºC in a temperature-controlled incubator on a 12 h light/dark cycle for total of 7 days. Ten RNA samples (Day 0: 2 conditions with 1 biological replicate each; Days 2 and 4: 2 conditions with 2 biological replicates each) were prepared for microarray hybridization. After total RNA extraction, rRNAs were removed and mRNA transcripts were amplified and labeled with Alexa Fluor 647. Two technical replicates were hybridized from each RNA sample. The microarray was designed based on selected Ruegeria pomeroyi DSS-3 genes and their orthologs in 12 other sequenced Roseobacter genomes. Probes were designed from the orthologs using the Hierarchical Probe Design (HPD) algorithm.

Project description:Purpose: Corals are major sources of dimethylsulphoniopropionate (DMSP), a compound that plays a central role in the global sulphur cycle. While DMSP biosynthesis pathways have been investigated in plants and algae, the molecular basis for its production by corals is unknown. Given its potential role as an osmolyte, the effect of salinity stress on levels of DMSP was investigated in both adults and juveniles (lacking photosynthetic symbionts) of the coral Acropora millepora. This study used transcriptomic data to analyse the effects of salinity over the coral A. millepora and to identify coral genes likely to be involved in DMSP biosynthesis. Methods: Adults coral transcriptomic libraries were constructed from samples exposed during 1 and 24 hours of salinity treatment (25 PSU) and control (35 PSU) conditions (n=5 per condition). Juveniles coral transcriptomic libraries were constructed from samples exposed to 24 and 48 hours of salinity treatment (28 PSU) and control (35 PSU) conditions (n=6 per condition). All libraries were sequenced by 100 bp paired-end in a HiSeq 2000. Reads were mapped onto the Acropora millepora genome using TopHat2 to produce a count data gene expression matrix for subsequent gene expression analysis using DESeq2 package. Results: In adult coral samples, 5.5 - 10.2 million RNAseq reads were obtained for each treatment sampling time while 3.4 - 8.8 million reads were obtained for each juvenile coral sample. The count matrix of the 26,622 A. millepora gene predictions were generated using htseq-count workflow. BlastP analysis of the A. millepora gene predictions led to the identification of coral members of gene families implicated in DMSP biosynthesis in other organisms, while RNA-seq data was used to identify the differentially expressed ones in response to hyposaline stress and on this basis were considered to be candidates for roles in DMSP biosynthesis in corals. Conclusions: Hyposaline stress increased DMSP production in both adults and aposymbiotic juvenile corals, and transcriptomic analyses highlighted the potential involvement of specific candidate genes in the production of DMSP via an alga-like pathway. The biochemistry of DMSP production is not well established for any eukaryotic system and, as the first animals in which it has been demonstrated, this is particularly true in the case of corals. Our RNA-seq results enabled the identification of candidates for roles in DMSP biosynthesis in corals but, given its critical roles in diverse biological processes, a thorough investigation of the molecular mechanisms leading to its production by corals is required.

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Bacterial lifestyle switch in response to algal metabolites

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