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Genome and Phenotype Microarray Analyses of Rhodococcus sp. BCP1 and Rhodococcus opacus R7: Genetic Determinants and Metabolic Abilities with Environmental Relevance.

Orro A, Cappelletti M, D'Ursi P, Milanesi L, Di Canito A, Zampolli J, Collina E, Decorosi F, Viti C, Fedi S, Presentato A, Zannoni D, Di Gennaro P - PLoS ONE (2015)

Bottom Line: Results show that R7 contains multiple genes for the degradation of a large set of aromatic and PAHs compounds, while a lower variability in terms of genes predicted to be involved in aromatic degradation was found in BCP1.According to this, in the BCP1 genome the smo gene cluster involved in the short-chain n-alkanes degradation, is included in one of the unique regions and it is not conserved in the Rhodococcus strains compared in this work.Data obtained underline the great potential of these two Rhodococcus spp. strains for biodegradation and environmental decontamination processes.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biomedical Technology, CNR, Segrate, Milano, Italy.

ABSTRACT
In this paper comparative genome and phenotype microarray analyses of Rhodococcus sp. BCP1 and Rhodococcus opacus R7 were performed. Rhodococcus sp. BCP1 was selected for its ability to grow on short-chain n-alkanes and R. opacus R7 was isolated for its ability to grow on naphthalene and on o-xylene. Results of genome comparison, including BCP1, R7, along with other Rhodococcus reference strains, showed that at least 30% of the genome of each strain presented unique sequences and only 50% of the predicted proteome was shared. To associate genomic features with metabolic capabilities of BCP1 and R7 strains, hundreds of different growth conditions were tested through Phenotype Microarray, by using Biolog plates and plates manually prepared with additional xenobiotic compounds. Around one-third of the surveyed carbon sources was utilized by both strains although R7 generally showed higher metabolic activity values compared to BCP1. Moreover, R7 showed broader range of nitrogen and sulphur sources. Phenotype Microarray data were combined with genomic analysis to genetically support the metabolic features of the two strains. The genome analysis allowed to identify some gene clusters involved in the metabolism of the main tested xenobiotic compounds. Results show that R7 contains multiple genes for the degradation of a large set of aromatic and PAHs compounds, while a lower variability in terms of genes predicted to be involved in aromatic degradation was found in BCP1. This genetic feature can be related to the strong genetic pressure exerted by the two different environment from which the two strains were isolated. According to this, in the BCP1 genome the smo gene cluster involved in the short-chain n-alkanes degradation, is included in one of the unique regions and it is not conserved in the Rhodococcus strains compared in this work. Data obtained underline the great potential of these two Rhodococcus spp. strains for biodegradation and environmental decontamination processes.

No MeSH data available.


Related in: MedlinePlus

Phenotype Microarray PM with different Nitrogen sources.Metabolic differences among R. opacus R7 and Rhodococcus sp. BCP1 in presence of different Nitrogen sources (AI, AII, AIII). Based on activity values of phenotype microarray analysis, threshold values were established for every plates. Determined thresholds were high (green), upper middle (light green), lower middle (orange) and low (red) for high, upper middle, lower middle and low activity, respectively.
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pone.0139467.g005: Phenotype Microarray PM with different Nitrogen sources.Metabolic differences among R. opacus R7 and Rhodococcus sp. BCP1 in presence of different Nitrogen sources (AI, AII, AIII). Based on activity values of phenotype microarray analysis, threshold values were established for every plates. Determined thresholds were high (green), upper middle (light green), lower middle (orange) and low (red) for high, upper middle, lower middle and low activity, respectively.

Mentions: Using the PM3 plate, the two Rhodococcus strains were tested for their ability to grow on 95 different nitrogen sources. R7 was able to utilize 74 out of 95 nitrogen sources, while BCP1 could grow only on 52 of the tested nitrogen compounds (Fig 5 Panel AI, AII, AIII). In particular, amongst the utilized N sources, R7 and BCP1 showed high metabolic activity on 50 and 24 compounds, respectively. Twenty-six N sources were specific for R7 strain growth, while 4 nitrogen sources were used by BCP1 and did not support the growth of R7. The nitrogen sources selective for BCP1 growth were D-lysine, DL-α-amino-caprylic acid, δ-amino-N-valeric acid, ε-amino-N-caproic acid. Interestingly, δ-amino-N-valeric acid was also a carbon source utilized by BCP1 but not by R7, suggesting the presence in BCP1 of transport and/or metabolic system specific for the utilization of this amino fatty acid compound. RAST annotation reported a number of genes predicted to be involved in amino acid metabolism that was double in R7 compared to BCP1 (Table 2). However, a similar number of genes predicted for nitrate and nitrite utilization and ammonia assimilation was annotated in BCP1 and R7 genomes (29 and 33 genes, respectively) (S5 Table). The higher activity values shown by BCP1 compared to R7 on ε-amino-N-caproic acid, D,L-α-amino-caprylic acid and δ-amino-N-valeric acid cannot be related to any genetic trait. However, some of the metabolic features of R7, not observed with BCP1, could be linked to the following genetic aspects. i) Considering the asparagine/aspartate metabolism, R7 has an asparaginase (EC 3.5.1.1) that converts asparagine in aspartate and an aspartate-ammonia lyase (EC 4.3.1.1) that transforms the aspartate in fumarate and ammonia that can be used as nitrogen source; the genes coding for the asparaginase and the aspartate-ammonia lyase are clustered together with a L-asparagine permease that is missing in BCP1 genome; both BCP1 and R7 have the asparagine synthetase (EC 6.3.5.4) that produces asparagine by ligating a NH3 group to aspartate using ATP; BCP1 genome lacks of an aspartic acid transporter. ii) Considering the arginine metabolism, R7 has genes coding for an arginase (EC 3.5.3.1) and an urease (EC 3.5.1.5) that convert arginine in urea and this in carbon dioxide and ammonia, respectively; R7 has also proton/glutamate aspartate symport protein that can mediate the import of glutamate and aspartate. iii) Considering the glutamine metabolism, the significantly higher activity of R7 on L-glutamine compared to BCP1 (at least double of BCP1 value) can be correlated with the presence of a L-glutaminase (EC 3.5.1.2) that hydrolyses L-glutamine in glutamate and ammonia. iv) considering the tyrosine metabolism, R7 has a tyramine:oxygen oxidoreductase (EC 1.4.-) which converts the tyramine that is included in tyrosine metabolism into 4-hydroxyphenylacetaldehyde and ammonia. This enzyme could also be related to the higher ability of R7 to utilize tyramine as carbon source.


Genome and Phenotype Microarray Analyses of Rhodococcus sp. BCP1 and Rhodococcus opacus R7: Genetic Determinants and Metabolic Abilities with Environmental Relevance.

Orro A, Cappelletti M, D'Ursi P, Milanesi L, Di Canito A, Zampolli J, Collina E, Decorosi F, Viti C, Fedi S, Presentato A, Zannoni D, Di Gennaro P - PLoS ONE (2015)

Phenotype Microarray PM with different Nitrogen sources.Metabolic differences among R. opacus R7 and Rhodococcus sp. BCP1 in presence of different Nitrogen sources (AI, AII, AIII). Based on activity values of phenotype microarray analysis, threshold values were established for every plates. Determined thresholds were high (green), upper middle (light green), lower middle (orange) and low (red) for high, upper middle, lower middle and low activity, respectively.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4591350&req=5

pone.0139467.g005: Phenotype Microarray PM with different Nitrogen sources.Metabolic differences among R. opacus R7 and Rhodococcus sp. BCP1 in presence of different Nitrogen sources (AI, AII, AIII). Based on activity values of phenotype microarray analysis, threshold values were established for every plates. Determined thresholds were high (green), upper middle (light green), lower middle (orange) and low (red) for high, upper middle, lower middle and low activity, respectively.
Mentions: Using the PM3 plate, the two Rhodococcus strains were tested for their ability to grow on 95 different nitrogen sources. R7 was able to utilize 74 out of 95 nitrogen sources, while BCP1 could grow only on 52 of the tested nitrogen compounds (Fig 5 Panel AI, AII, AIII). In particular, amongst the utilized N sources, R7 and BCP1 showed high metabolic activity on 50 and 24 compounds, respectively. Twenty-six N sources were specific for R7 strain growth, while 4 nitrogen sources were used by BCP1 and did not support the growth of R7. The nitrogen sources selective for BCP1 growth were D-lysine, DL-α-amino-caprylic acid, δ-amino-N-valeric acid, ε-amino-N-caproic acid. Interestingly, δ-amino-N-valeric acid was also a carbon source utilized by BCP1 but not by R7, suggesting the presence in BCP1 of transport and/or metabolic system specific for the utilization of this amino fatty acid compound. RAST annotation reported a number of genes predicted to be involved in amino acid metabolism that was double in R7 compared to BCP1 (Table 2). However, a similar number of genes predicted for nitrate and nitrite utilization and ammonia assimilation was annotated in BCP1 and R7 genomes (29 and 33 genes, respectively) (S5 Table). The higher activity values shown by BCP1 compared to R7 on ε-amino-N-caproic acid, D,L-α-amino-caprylic acid and δ-amino-N-valeric acid cannot be related to any genetic trait. However, some of the metabolic features of R7, not observed with BCP1, could be linked to the following genetic aspects. i) Considering the asparagine/aspartate metabolism, R7 has an asparaginase (EC 3.5.1.1) that converts asparagine in aspartate and an aspartate-ammonia lyase (EC 4.3.1.1) that transforms the aspartate in fumarate and ammonia that can be used as nitrogen source; the genes coding for the asparaginase and the aspartate-ammonia lyase are clustered together with a L-asparagine permease that is missing in BCP1 genome; both BCP1 and R7 have the asparagine synthetase (EC 6.3.5.4) that produces asparagine by ligating a NH3 group to aspartate using ATP; BCP1 genome lacks of an aspartic acid transporter. ii) Considering the arginine metabolism, R7 has genes coding for an arginase (EC 3.5.3.1) and an urease (EC 3.5.1.5) that convert arginine in urea and this in carbon dioxide and ammonia, respectively; R7 has also proton/glutamate aspartate symport protein that can mediate the import of glutamate and aspartate. iii) Considering the glutamine metabolism, the significantly higher activity of R7 on L-glutamine compared to BCP1 (at least double of BCP1 value) can be correlated with the presence of a L-glutaminase (EC 3.5.1.2) that hydrolyses L-glutamine in glutamate and ammonia. iv) considering the tyrosine metabolism, R7 has a tyramine:oxygen oxidoreductase (EC 1.4.-) which converts the tyramine that is included in tyrosine metabolism into 4-hydroxyphenylacetaldehyde and ammonia. This enzyme could also be related to the higher ability of R7 to utilize tyramine as carbon source.

Bottom Line: Results show that R7 contains multiple genes for the degradation of a large set of aromatic and PAHs compounds, while a lower variability in terms of genes predicted to be involved in aromatic degradation was found in BCP1.According to this, in the BCP1 genome the smo gene cluster involved in the short-chain n-alkanes degradation, is included in one of the unique regions and it is not conserved in the Rhodococcus strains compared in this work.Data obtained underline the great potential of these two Rhodococcus spp. strains for biodegradation and environmental decontamination processes.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biomedical Technology, CNR, Segrate, Milano, Italy.

ABSTRACT
In this paper comparative genome and phenotype microarray analyses of Rhodococcus sp. BCP1 and Rhodococcus opacus R7 were performed. Rhodococcus sp. BCP1 was selected for its ability to grow on short-chain n-alkanes and R. opacus R7 was isolated for its ability to grow on naphthalene and on o-xylene. Results of genome comparison, including BCP1, R7, along with other Rhodococcus reference strains, showed that at least 30% of the genome of each strain presented unique sequences and only 50% of the predicted proteome was shared. To associate genomic features with metabolic capabilities of BCP1 and R7 strains, hundreds of different growth conditions were tested through Phenotype Microarray, by using Biolog plates and plates manually prepared with additional xenobiotic compounds. Around one-third of the surveyed carbon sources was utilized by both strains although R7 generally showed higher metabolic activity values compared to BCP1. Moreover, R7 showed broader range of nitrogen and sulphur sources. Phenotype Microarray data were combined with genomic analysis to genetically support the metabolic features of the two strains. The genome analysis allowed to identify some gene clusters involved in the metabolism of the main tested xenobiotic compounds. Results show that R7 contains multiple genes for the degradation of a large set of aromatic and PAHs compounds, while a lower variability in terms of genes predicted to be involved in aromatic degradation was found in BCP1. This genetic feature can be related to the strong genetic pressure exerted by the two different environment from which the two strains were isolated. According to this, in the BCP1 genome the smo gene cluster involved in the short-chain n-alkanes degradation, is included in one of the unique regions and it is not conserved in the Rhodococcus strains compared in this work. Data obtained underline the great potential of these two Rhodococcus spp. strains for biodegradation and environmental decontamination processes.

No MeSH data available.


Related in: MedlinePlus