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Application of a Novel "Pan-Genome"-Based Strategy for Assigning RNAseq Transcript Reads to Staphylococcus aureus Strains.

Chaves-Moreno D, Wos-Oxley ML, Jáuregui R, Medina E, Oxley AP, Pieper DH - PLoS ONE (2015)

Bottom Line: The pan-genome of S. aureus and its associated core and accessory components were compiled based on 25 genomes and comprises a total of 65,557 proteins clustering into 4,198 Orthologous Groups (OGs).The OG database generated in this study represents a useful tool to obtain a snapshot of the functional attributes of S. aureus under different in vitro and in vivo conditions.The approach proved to be advantageous to assign sequencing reads to bacterial strains when RNAseq data is derived from samples where strain information and/or the corresponding genome/s are unavailable.

View Article: PubMed Central - PubMed

Affiliation: Microbial Interactions and Processes Research Group, Helmholtz Centre for Infection Research, Braunschweig, Germany.

ABSTRACT
Understanding the behaviour of opportunistic pathogens such as Staphylococcus aureus in their natural human niche holds great medical interest. With the development of sensitive molecular methods and deep-sequencing technology, it is now possible to robustly assess the global transcriptome of bacterial species in their human habitat. However, as the genomes of the colonizing strains are often not available compiling the pan-genome for the species of interest may provide an effective method to reliably and rapidly compile the transcriptome of a bacterial species. The pan-genome of S. aureus and its associated core and accessory components were compiled based on 25 genomes and comprises a total of 65,557 proteins clustering into 4,198 Orthologous Groups (OGs). The generated gene catalogue was used to assign RNAseq-derived sequence reads to S. aureus in a variety of in vitro and in vivo samples. In all cases, the number of reads that could be assigned to S. aureus was greater using the OG database than using a reference genome. Growth of two S. aureus strains in synthetic nasal medium confirmed that both strains experienced strong iron starvation. Traits such as purine metabolism appeared to be more affected in a typical nasal colonizer than in a strain representative of the S. aureus USA300 lineage. Mapping sequencing reads from a metatranscriptome generated from the human anterior nares allowed the identification of genes highly expressed by S. aureus in vivo. The OG database generated in this study represents a useful tool to obtain a snapshot of the functional attributes of S. aureus under different in vitro and in vivo conditions. The approach proved to be advantageous to assign sequencing reads to bacterial strains when RNAseq data is derived from samples where strain information and/or the corresponding genome/s are unavailable.

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Comparison of the global S. aureus transcriptome under in vivo and in vitro growth conditions.Comparison of the global S. aureus transcriptomic profile from 7 RNAseq libraries using (A) Principal Coordinate Analysis (PCoA) and (B) agglomerative hierachical clustering (group-average linkage). Conditions included: in vitro exponential and stationary phase cultures of S. aureus USA300 strain LAC grown in Brain Heart Infusion (BHI) media; in vitro EX and ST phase cultures of S. aureus strain IPL32 grown in BHI; in vitro EX phase cultures of both strains grown in a Synthetic Nasal Medium (SNM) that mimics the composition of the human nasal secretions [17]; and in vivo S. aureus within the complex bacterial community of the human anterior nares. The Bray-curtis similarity algorithm was used to assess the similarity between samples based on the relative abundance of transcripts for each OG, where 70.6% of the total variation could be explained by PCO1 and PCO2.
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pone.0145861.g002: Comparison of the global S. aureus transcriptome under in vivo and in vitro growth conditions.Comparison of the global S. aureus transcriptomic profile from 7 RNAseq libraries using (A) Principal Coordinate Analysis (PCoA) and (B) agglomerative hierachical clustering (group-average linkage). Conditions included: in vitro exponential and stationary phase cultures of S. aureus USA300 strain LAC grown in Brain Heart Infusion (BHI) media; in vitro EX and ST phase cultures of S. aureus strain IPL32 grown in BHI; in vitro EX phase cultures of both strains grown in a Synthetic Nasal Medium (SNM) that mimics the composition of the human nasal secretions [17]; and in vivo S. aureus within the complex bacterial community of the human anterior nares. The Bray-curtis similarity algorithm was used to assess the similarity between samples based on the relative abundance of transcripts for each OG, where 70.6% of the total variation could be explained by PCO1 and PCO2.

Mentions: A data matrix comparing the assigned reads counted against the OG database (S2 Table) was subsequently compiled for each of the 7 conditions and the different conditions were compared using group-average agglomerative hierarchical clustering and Principal Co-ordinate Analysis (PcoA) (Fig 2). The global transcription profiles of the strains USA300 LAC and IPL32 grown under identical in vitro conditions always clustered tightly together, sharing roughly 80% similarity within each of the 3 in vitro growth conditions. Furthermore, all transcriptomes from the 6 in vitro samples, irrespective of strain, media or growth phase differences shared a global similarity of ≥ 67%. However, the transcriptome of S. aureus colonizing the human anterior nares shared on average only 47% similarity with any of the transcriptomes from S. aureus growing under different in vitro conditions. This highlights the strong influence of the environmental conditions encountered within the host in the adaptive transcriptional response of S. aureus.


Application of a Novel "Pan-Genome"-Based Strategy for Assigning RNAseq Transcript Reads to Staphylococcus aureus Strains.

Chaves-Moreno D, Wos-Oxley ML, Jáuregui R, Medina E, Oxley AP, Pieper DH - PLoS ONE (2015)

Comparison of the global S. aureus transcriptome under in vivo and in vitro growth conditions.Comparison of the global S. aureus transcriptomic profile from 7 RNAseq libraries using (A) Principal Coordinate Analysis (PCoA) and (B) agglomerative hierachical clustering (group-average linkage). Conditions included: in vitro exponential and stationary phase cultures of S. aureus USA300 strain LAC grown in Brain Heart Infusion (BHI) media; in vitro EX and ST phase cultures of S. aureus strain IPL32 grown in BHI; in vitro EX phase cultures of both strains grown in a Synthetic Nasal Medium (SNM) that mimics the composition of the human nasal secretions [17]; and in vivo S. aureus within the complex bacterial community of the human anterior nares. The Bray-curtis similarity algorithm was used to assess the similarity between samples based on the relative abundance of transcripts for each OG, where 70.6% of the total variation could be explained by PCO1 and PCO2.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4696825&req=5

pone.0145861.g002: Comparison of the global S. aureus transcriptome under in vivo and in vitro growth conditions.Comparison of the global S. aureus transcriptomic profile from 7 RNAseq libraries using (A) Principal Coordinate Analysis (PCoA) and (B) agglomerative hierachical clustering (group-average linkage). Conditions included: in vitro exponential and stationary phase cultures of S. aureus USA300 strain LAC grown in Brain Heart Infusion (BHI) media; in vitro EX and ST phase cultures of S. aureus strain IPL32 grown in BHI; in vitro EX phase cultures of both strains grown in a Synthetic Nasal Medium (SNM) that mimics the composition of the human nasal secretions [17]; and in vivo S. aureus within the complex bacterial community of the human anterior nares. The Bray-curtis similarity algorithm was used to assess the similarity between samples based on the relative abundance of transcripts for each OG, where 70.6% of the total variation could be explained by PCO1 and PCO2.
Mentions: A data matrix comparing the assigned reads counted against the OG database (S2 Table) was subsequently compiled for each of the 7 conditions and the different conditions were compared using group-average agglomerative hierarchical clustering and Principal Co-ordinate Analysis (PcoA) (Fig 2). The global transcription profiles of the strains USA300 LAC and IPL32 grown under identical in vitro conditions always clustered tightly together, sharing roughly 80% similarity within each of the 3 in vitro growth conditions. Furthermore, all transcriptomes from the 6 in vitro samples, irrespective of strain, media or growth phase differences shared a global similarity of ≥ 67%. However, the transcriptome of S. aureus colonizing the human anterior nares shared on average only 47% similarity with any of the transcriptomes from S. aureus growing under different in vitro conditions. This highlights the strong influence of the environmental conditions encountered within the host in the adaptive transcriptional response of S. aureus.

Bottom Line: The pan-genome of S. aureus and its associated core and accessory components were compiled based on 25 genomes and comprises a total of 65,557 proteins clustering into 4,198 Orthologous Groups (OGs).The OG database generated in this study represents a useful tool to obtain a snapshot of the functional attributes of S. aureus under different in vitro and in vivo conditions.The approach proved to be advantageous to assign sequencing reads to bacterial strains when RNAseq data is derived from samples where strain information and/or the corresponding genome/s are unavailable.

View Article: PubMed Central - PubMed

Affiliation: Microbial Interactions and Processes Research Group, Helmholtz Centre for Infection Research, Braunschweig, Germany.

ABSTRACT
Understanding the behaviour of opportunistic pathogens such as Staphylococcus aureus in their natural human niche holds great medical interest. With the development of sensitive molecular methods and deep-sequencing technology, it is now possible to robustly assess the global transcriptome of bacterial species in their human habitat. However, as the genomes of the colonizing strains are often not available compiling the pan-genome for the species of interest may provide an effective method to reliably and rapidly compile the transcriptome of a bacterial species. The pan-genome of S. aureus and its associated core and accessory components were compiled based on 25 genomes and comprises a total of 65,557 proteins clustering into 4,198 Orthologous Groups (OGs). The generated gene catalogue was used to assign RNAseq-derived sequence reads to S. aureus in a variety of in vitro and in vivo samples. In all cases, the number of reads that could be assigned to S. aureus was greater using the OG database than using a reference genome. Growth of two S. aureus strains in synthetic nasal medium confirmed that both strains experienced strong iron starvation. Traits such as purine metabolism appeared to be more affected in a typical nasal colonizer than in a strain representative of the S. aureus USA300 lineage. Mapping sequencing reads from a metatranscriptome generated from the human anterior nares allowed the identification of genes highly expressed by S. aureus in vivo. The OG database generated in this study represents a useful tool to obtain a snapshot of the functional attributes of S. aureus under different in vitro and in vivo conditions. The approach proved to be advantageous to assign sequencing reads to bacterial strains when RNAseq data is derived from samples where strain information and/or the corresponding genome/s are unavailable.

Show MeSH
Related in: MedlinePlus