A five-species transcriptome array for oral mixed-biofilm studies
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PROVIDER: PRJNA140619 | ENA |
REPOSITORIES: ENA
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Project description:Investigation of whole genome gene expression levels of P. gingivalis W83, F. nucleatum DSMZ 25586, S. sanguinis SK36, A. actinomycetemcomitans HK1651, S. mutans UA159 in an 24 h old culture. Additionally, whole genome gene expression level changes of S. mutans UA159 biofilm cells after co-cultivation with S. mitis ATCC 11843 were compared to its single species biofilm growth after 24 h. Aim: Demonstration of the usefulness of a five-species gene expression array. Multiple probes per gene enabled identification of single inter-species cross-hybridizing probes. The deletion of such probes lead almost not to the deletion of the whole gene. This was investigated and confirmed by a two-species biofilm expression analysis: The here described array was used for the identification of genes of S. mutans influenced by the presence of S. mitis. Materials and Methods: P. gingivalis W83, F. nucleatum DSMZ 25586, S. sanguinis SK36, A. actinomycetemcomitans HK1651,and S. mutans UA159 were grown in CDM/succrose or artificial saliva/galactose in a single-species culture for 24 h anaerobically resulting in biofilm structures or monolayers. Total RNA was isolated and used for microarray analysis. Probes were analysed for the presence of biological false positive signals caused by cross-hybridizing probes of one of the other species presented on the chip. Further, a simple procedure was developed for automatical identification and deletion of false positive signals caused by washing artefacts, resulting in a more reliable outcome. In the case of the S. mutans/S. mitis mixed-species biofilm, both species were cultured together for 24 h like previously described. The found gene regulations were verified by RT-PCR. Results: Experiments with cDNA from 24 h old single-species cultures allowed the identification of cross-species hybridizing probes on the array, which can be eliminated in mixed-species experimental settings without the need to exclude the whole genes from the analysis. Between 69 % and almost 100 % represented genomes on this array were found actively transcribed under the mono-species monolayer and biofilm conditions used here. S. mutans / S. mitis co-culture: Physiological investigations revealed an increase in S. mutans biofilm mass with a decrease in pH-value under the influence of S. mitis, thereby confirming previously published data. A stringent fold change cut-off of 2 (p<0.05) identified 19 S. mutans transcripts with increased abundance, and 11 with decreased abundance compared to a S. mutans mono-species biofilm. Many of the genes have previously been found differentially regulated under general and acid stress, thereby confirming the value of this array. Conclusions: Taken together, this new array allows transcriptome studies on multi-species oral biofilm interactions and could become an important asset in future oral biofilm and inhibitor/therapy studies.
2011-12-09 | GSE28841 | GEO
Project description:Investigation of whole genome gene expression levels of P. gingivalis W83, F. nucleatum DSMZ 25586, S. sanguinis SK36, A. actinomycetemcomitans HK1651, S. mutans UA159 in an 24 h old culture. Additionally, whole genome gene expression level changes of S. mutans UA159 biofilm cells after co-cultivation with S. mitis ATCC 11843 were compared to its single species biofilm growth after 24 h. Aim: Demonstration of the usefulness of a five-species gene expression array. Multiple probes per gene enabled identification of single inter-species cross-hybridizing probes. The deletion of such probes lead almost not to the deletion of the whole gene. This was investigated and confirmed by a two-species biofilm expression analysis: The here described array was used for the identification of genes of S. mutans influenced by the presence of S. mitis. Materials and Methods: P. gingivalis W83, F. nucleatum DSMZ 25586, S. sanguinis SK36, A. actinomycetemcomitans HK1651,and S. mutans UA159 were grown in CDM/succrose or artificial saliva/galactose in a single-species culture for 24 h anaerobically resulting in biofilm structures or monolayers. Total RNA was isolated and used for microarray analysis. Probes were analysed for the presence of biological false positive signals caused by cross-hybridizing probes of one of the other species presented on the chip. Further, a simple procedure was developed for automatical identification and deletion of false positive signals caused by washing artefacts, resulting in a more reliable outcome. In the case of the S. mutans/S. mitis mixed-species biofilm, both species were cultured together for 24 h like previously described. The found gene regulations were verified by RT-PCR. Results: Experiments with cDNA from 24 h old single-species cultures allowed the identification of cross-species hybridizing probes on the array, which can be eliminated in mixed-species experimental settings without the need to exclude the whole genes from the analysis. Between 69 % and almost 100 % represented genomes on this array were found actively transcribed under the mono-species monolayer and biofilm conditions used here. S. mutans / S. mitis co-culture: Physiological investigations revealed an increase in S. mutans biofilm mass with a decrease in pH-value under the influence of S. mitis, thereby confirming previously published data. A stringent fold change cut-off of 2 (p<0.05) identified 19 S. mutans transcripts with increased abundance, and 11 with decreased abundance compared to a S. mutans mono-species biofilm. Many of the genes have previously been found differentially regulated under general and acid stress, thereby confirming the value of this array. Conclusions: Taken together, this new array allows transcriptome studies on multi-species oral biofilm interactions and could become an important asset in future oral biofilm and inhibitor/therapy studies. The chip study used pooled total RNA recovered from three biologically independent mono-species biofilms or adherent cells/monolayers of P. gingivalis W83, F. nucleatum DSMZ 25586, S. sanguinis SK36, A. actinomycetemcomitans HK1651, and S. mutans UA159. In the case of gene expression analysis of S. mutans/S.mitis biofilm structures compared to the single species biofilm of S. mutans three separate single and three separate two-species biofilm cultures were analysed. Each chip measured the expression level of all together 10186 genes (1883 genes of P. gingivalis W83, 1964 genes of F. nucleatum DSMZ 25586, 2244 genes of S. sanguinis SK36, 2168 genes of A. actinomycetemcomitans HK1651, 1927 genes of S. mutans UA159) with up to thirteen 60-mer probes per gene and with a three-fold technical redundancy.
2011-12-09 | E-GEOD-28841 | biostudies-arrayexpress
Project description:BackgroundOral polymicrobial interactions and biofilm formation are associated with initiation and progression of caries, gingivitis, and periodontitis. Transcriptome studies of such interactions, allowing a first mechanistic insight, are hampered by current single-species array designs.Methodology/principal findingsIn this study we used 385 K NimbleGene™ technology for design and evaluation of an array covering the full genomes of 5 important physiological-, cariogenic-, and periodontitis-associated microorganisms (Streptococcus sanguinis, Streptococcus mutans, Fusobacterium nucleatum, Aggregatibacter actinomycetemcomitans, and Porphyromonas gingivalis). Array hybridization was done with cDNA from cultures grown for 24 h anaerobically. Single species experiments identified cross-species hybridizing array probes. These probes could be neglected in a mixed-species experimental setting without the need to exclude the whole genes from the analysis. Between 69% and almost 99% of the genomes were actively transcribed under the mono-species planktonic, monolayer, and biofilm conditions. The influence of Streptococcus mitis (not represented on the array) on S. mutans gene transcription was determined as a test for a dual-species mixed biofilm setup. Phenotypically, under the influence of S. mitis an increase in S. mutans biofilm mass and a decrease in media pH-value were noticed, thereby confirming previously published data. Employing a stringent cut-off (2-fold, p<0.05), 19 S. mutans transcripts were identified with increased abundance, and 11 with decreased abundance compared to a S. mutans mono-species biofilm. Several of these genes have previously been found differentially regulated under general and acid stress, thereby confirming the value of this array.Conclusions/significanceThis new array allows transcriptome studies on multi-species oral biofilm interactions. It may become an important asset in future oral biofilm and inhibitor/therapy studies.
| S-EPMC3237422 | biostudies-literature
Project description:Transcription profile of Escherichia coli cells in mono-species pure biofilms was compared to that of E. coli cells in E. coli-Stenotrophomonas maltophilia dual-species biofilms. E. coli cells were separated from dual-species biofilms before total RNA extraction to eliminate possible cross hybridization from S. maltophilia transcripts. The separation method was developed by combining the use of reagent RNAlater and immuno-magnetic separation. Pure E. coli biofilms were processed with the same separation protocol before RNA extraction. Two condition experiments: E. coli mono-species biofilm vs E. coli in mixed-species biofilm. Two biological replicates with independently grown and harvested biofilms. Each biological replicate has two or three technical replicates of hybridization on microarray slides. Each slide has three built-in replicates for each probe.
2013-12-30 | E-GEOD-24912 | biostudies-arrayexpress
Project description:To explain enhanced biofilm formation and increased dissemination of S. epidermidis in mixed-species biofilms, microarrays were used to explore differential gene expression of S. epidermidis in mixed-species biofilms. One sample from single species biofilm (S1) and mixed-species biofilm (SC2) were excluded from analyses for outliers. We observed upregulation (2.7%) and down regulation (6%) of S. epidermidis genes in mixed-species biofilms. Autolysis repressors lrgA and lrgB were down regulated 36-fold and 27-fold respectively and was associated with increased eDNA possibly due to enhanced autolysis in mixed-species biofilms. These data suggest that bacterial autolysis and release of eDNA in the biofilm matrix may be responsible for enhancement and dissemination of mixed-species biofilms of S. epidermidis and C. albicans.
2012-02-28 | GSE35438 | GEO
Project description:To explain enhanced biofilm formation and increased dissemination of S. epidermidis in mixed-species biofilms, microarrays were used to explore differential gene expression of S. epidermidis in mixed-species biofilms. One sample from single species biofilm (S1) and mixed-species biofilm (SC2) were excluded from analyses for outliers. We observed upregulation (2.7%) and down regulation (6%) of S. epidermidis genes in mixed-species biofilms. Autolysis repressors lrgA and lrgB were down regulated 36-fold and 27-fold respectively and was associated with increased eDNA possibly due to enhanced autolysis in mixed-species biofilms. These data suggest that bacterial autolysis and release of eDNA in the biofilm matrix may be responsible for enhancement and dissemination of mixed-species biofilms of S. epidermidis and C. albicans. Staphylococcal gene expression in mixed-species biofilms with Candida and in single species biofilms of S. epidermidis were analyzed. The experiment was repeated thrice on 3 different days (3 biological replicates each for single species biofilms of S. epidermidis and mixed-species biofilms). Only 2 biological replicates were analyzed and one biological replicate was not analyzed (S1 and SC1 - raw data files are provided on the Series record). Single species biofilms of S. epidermidis (strain 1457) and C. albicans (strain 32354) and mixed-species biofilms were formed on 6-well tissue culture plates. Five ml of organism suspensions (O.D. 0.3, S. epidermidis 107 CFU/ml or C. albicans 105 CFU/ml) or 2.5 ml each for mixed-species biofilms for 24 hr. RNA was harvested from single species and mixed-species biofilms.
2012-02-28 | E-GEOD-35438 | biostudies-arrayexpress
Project description:Transcriptome analysis to determine the impact of oral exposure (in a sugar meal) to the liquid supernatant (i.e. LB culture media) of Chromobacterium sp. Panama biofilm culture. The biofilm supernatant (i.e. media) was first filtered with a 0.2uM filter to remove all live bacterial cells. It was then mixed with 10% sucrose, and a control sucrose meal was mixed with filtered LB. Mosquitoes were exposed to each sugar meal for 24 hours and then midguts were dissected from 20 adult females per treatment. The entire experiment was performed 4 independent times.
2018-03-31 | GSE109727 | GEO
Project description:Anode-associated multi-species exoelectrogenic biofilms are essential to the function of bioelectrochemical systems (BESs). The investigation of electrode-associated biofilms is critical to advance understanding of the function of individual members within communities that thrive using an electrode as the terminal electron acceptor. This study focusses on the analysis of a model biofilm community consisting of Shewanella oneidensis, Geobacter sulfurreducens and Geobacter metallireducens. The conducted experiments revealed that the organisms can build a stable biofilm on an electrode surface that is rather resilient to changes in the redox potential of the anode surface. The community operated at maximum electron transfer rates with electrode potentials of 0.04 V versus normal hydrogen electrode. Current densities decreased gradually with lower potentials and reached half-maximal values at -0.08 V. A positive interaction of the individual strains could be observed in our experiments. At least S. oneidensis and G. sulfurreducens show an upregulation of their central metabolism as a response to cultivation under mixed-species conditions. Interestingly, G. sulfurreducens was detected in the planktonic phase of the bioelectrochemical reactors only in mixed-culture experiments but not when it was grown in the absence of the other two organisms. It is possible that G. sulfurreducens cells used flavins which were released by S. oneidensis cells as electron shuttles. This would allow the organism to broaden its environmental niche. To the best of our knowledge, this is the first study describing the dynamics of biofilm formation of a model exoelectrogenic community, the resilience of the biofilm, and the molecular responses towards mixed-species conditions.
2017-01-18 | GSE79750 | GEO
Project description:Transcriptome profiling of five mixed tissues for juvenile Eriocheir sinensis
| PRJNA757979 | ENA