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Characterization of the algC gene expression pattern in the multidrug resistant Acinetobacter baumannii AIIMS 7 and correlation with biofilm development on abiotic surface.

Sahu PK, Iyer PS, Barage SH, Sonawane KD, Chopade BA - ScientificWorldJournal (2014)

Bottom Line: Comparison revealed differential algC expression pattern with maximum 81.59-fold increase in biofilm cells versus 3.24-fold in planktonic cells (P < 0.05).Expression levels strongly correlated with specific biofilm stages (scale of 3 to 96 h), coinciding maximum at initial surface attachment stage (9 h) and biofilm maturation stage (48 h).Moreover, molecular dynamics analysis on the three-dimensional structure of PMM/PGM (simulated up to 10 ns) revealed enzyme structure as stable and similar to that in P. aeruginosa (synthesis of alginate and lipopolysaccharide core) and involved in constitution of biofilm EPS (extracellular polymeric substances).

View Article: PubMed Central - PubMed

Affiliation: Institute of Bioinformatics and Biotechnology, University of Pune, Pune 411 007, India ; Ispat General Hospital, SAIL, Rourkela 769 005, India.

ABSTRACT
Relative quantification of algC gene expression was evaluated in the multidrug resistant strain Acinetobacter baumannii AIIMS 7 biofilm (3 to 96 h, on polystyrene surface) compared to the planktonic counterparts. Comparison revealed differential algC expression pattern with maximum 81.59-fold increase in biofilm cells versus 3.24-fold in planktonic cells (P < 0.05). Expression levels strongly correlated with specific biofilm stages (scale of 3 to 96 h), coinciding maximum at initial surface attachment stage (9 h) and biofilm maturation stage (48 h). Cloning, heterologous expression, and bioinformatics analyses indicated algC gene product as the bifunctional enzyme phosphomannomutase/phosphoglucomutase (PMM/PGM) of ∼ 53 kDa size, which augmented biofilms significantly in algC clones compared to controls (lacking algC gene), further localized by scanning electron microscopy. Moreover, molecular dynamics analysis on the three-dimensional structure of PMM/PGM (simulated up to 10 ns) revealed enzyme structure as stable and similar to that in P. aeruginosa (synthesis of alginate and lipopolysaccharide core) and involved in constitution of biofilm EPS (extracellular polymeric substances). Our observation on differential expression pattern of algC having strong correlation with important biofilm stages, scanning electron-microscopic evidence of biofilm augmentation taken together with predictive enzyme functions via molecular dynamic (MD) simulation, proposes a new basis of A. baumannii AIIMS 7 biofilm development on inanimate surfaces.

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(a)–(j) Microscopic visualization of A. baumannii AIIMS 7 biofilms. Representative bright field micrographs depicting static biofilms formed on polystyrene microtitre surface at respective time points (3, 6, 9, 12, 18, 24, 36, 48, 72, 96 h) (magnification, 100x).
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fig3: (a)–(j) Microscopic visualization of A. baumannii AIIMS 7 biofilms. Representative bright field micrographs depicting static biofilms formed on polystyrene microtitre surface at respective time points (3, 6, 9, 12, 18, 24, 36, 48, 72, 96 h) (magnification, 100x).

Mentions: To visualize the growth pattern of biofilm, bright-field microscopy was performed on biofilms growing on polystyrene surface, which showed varied biofilm morphology at gradual time points (Figure 3) with a strong concurrence with quantitative evaluation of biofilm growth pattern (Figure 2, as described above). At initial attachment stages, planktonic cells were seen scantily aggregated (Figures 3(a) and 3(b)) but gradually the micro-aggregation on the surface increased, subsequently making completely attached micro-colonies at 9 h which could be seen clearly (Figure 3(c)). Further stages of growth (12, 18, and 24 h) showed consolidation of micro-colonies leading to stable structures of biofilm (Figures 3(d)–3(f)). At 36–48 h, fully mature biofilm communities of A. baumannii were seen, enriched with thick EPS matrices (Figures 3(g) and 3(h)), followed by appearance of robust three-dimensional biofilm structures, which were found persistent till 72 and 96 h (Figures 3(i) and 3(j)). Overall, visualization of biofilm growth at developing stages was in tandem with the patterns of biofilm growth as well as algC gene expression.


Characterization of the algC gene expression pattern in the multidrug resistant Acinetobacter baumannii AIIMS 7 and correlation with biofilm development on abiotic surface.

Sahu PK, Iyer PS, Barage SH, Sonawane KD, Chopade BA - ScientificWorldJournal (2014)

(a)–(j) Microscopic visualization of A. baumannii AIIMS 7 biofilms. Representative bright field micrographs depicting static biofilms formed on polystyrene microtitre surface at respective time points (3, 6, 9, 12, 18, 24, 36, 48, 72, 96 h) (magnification, 100x).
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4269089&req=5

fig3: (a)–(j) Microscopic visualization of A. baumannii AIIMS 7 biofilms. Representative bright field micrographs depicting static biofilms formed on polystyrene microtitre surface at respective time points (3, 6, 9, 12, 18, 24, 36, 48, 72, 96 h) (magnification, 100x).
Mentions: To visualize the growth pattern of biofilm, bright-field microscopy was performed on biofilms growing on polystyrene surface, which showed varied biofilm morphology at gradual time points (Figure 3) with a strong concurrence with quantitative evaluation of biofilm growth pattern (Figure 2, as described above). At initial attachment stages, planktonic cells were seen scantily aggregated (Figures 3(a) and 3(b)) but gradually the micro-aggregation on the surface increased, subsequently making completely attached micro-colonies at 9 h which could be seen clearly (Figure 3(c)). Further stages of growth (12, 18, and 24 h) showed consolidation of micro-colonies leading to stable structures of biofilm (Figures 3(d)–3(f)). At 36–48 h, fully mature biofilm communities of A. baumannii were seen, enriched with thick EPS matrices (Figures 3(g) and 3(h)), followed by appearance of robust three-dimensional biofilm structures, which were found persistent till 72 and 96 h (Figures 3(i) and 3(j)). Overall, visualization of biofilm growth at developing stages was in tandem with the patterns of biofilm growth as well as algC gene expression.

Bottom Line: Comparison revealed differential algC expression pattern with maximum 81.59-fold increase in biofilm cells versus 3.24-fold in planktonic cells (P < 0.05).Expression levels strongly correlated with specific biofilm stages (scale of 3 to 96 h), coinciding maximum at initial surface attachment stage (9 h) and biofilm maturation stage (48 h).Moreover, molecular dynamics analysis on the three-dimensional structure of PMM/PGM (simulated up to 10 ns) revealed enzyme structure as stable and similar to that in P. aeruginosa (synthesis of alginate and lipopolysaccharide core) and involved in constitution of biofilm EPS (extracellular polymeric substances).

View Article: PubMed Central - PubMed

Affiliation: Institute of Bioinformatics and Biotechnology, University of Pune, Pune 411 007, India ; Ispat General Hospital, SAIL, Rourkela 769 005, India.

ABSTRACT
Relative quantification of algC gene expression was evaluated in the multidrug resistant strain Acinetobacter baumannii AIIMS 7 biofilm (3 to 96 h, on polystyrene surface) compared to the planktonic counterparts. Comparison revealed differential algC expression pattern with maximum 81.59-fold increase in biofilm cells versus 3.24-fold in planktonic cells (P < 0.05). Expression levels strongly correlated with specific biofilm stages (scale of 3 to 96 h), coinciding maximum at initial surface attachment stage (9 h) and biofilm maturation stage (48 h). Cloning, heterologous expression, and bioinformatics analyses indicated algC gene product as the bifunctional enzyme phosphomannomutase/phosphoglucomutase (PMM/PGM) of ∼ 53 kDa size, which augmented biofilms significantly in algC clones compared to controls (lacking algC gene), further localized by scanning electron microscopy. Moreover, molecular dynamics analysis on the three-dimensional structure of PMM/PGM (simulated up to 10 ns) revealed enzyme structure as stable and similar to that in P. aeruginosa (synthesis of alginate and lipopolysaccharide core) and involved in constitution of biofilm EPS (extracellular polymeric substances). Our observation on differential expression pattern of algC having strong correlation with important biofilm stages, scanning electron-microscopic evidence of biofilm augmentation taken together with predictive enzyme functions via molecular dynamic (MD) simulation, proposes a new basis of A. baumannii AIIMS 7 biofilm development on inanimate surfaces.

Show MeSH
Related in: MedlinePlus