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Phenotypic and Genomic Properties of Chitinispirillum alkaliphilum gen. nov., sp. nov., A Haloalkaliphilic Anaerobic Chitinolytic Bacterium Representing a Novel Class in the Phylum Fibrobacteres.

Sorokin DY, Rakitin AL, Gumerov VM, Beletsky AV, Sinninghe Damsté JS, Mardanov AV, Ravin NV - Front Microbiol (2016)

Bottom Line: The chitinolytic activity was associated with cells.Analysis of the 4.4 Mb draft genome identified pathways for chitin utilization, particularly, secreted chitinases linked to the cell surface, as well as genes for the hydrolysis of other polysaccharides and fermentation of sugars, while the genes needed for aerobic and anaerobic respiration were absent.The phylogenetic analysis using 16S rRNA gene and ribosomal proteins indicated that ACht6-1 forms a novel deep lineage at the class level within the bacterial candidate division TG3.

View Article: PubMed Central - PubMed

Affiliation: Winogradsky Institute of Microbiology, Research Center of Biotechnology of the Russian Academy of SciencesMoscow, Russia; Department of Biotechnology, Delft University of TechnologyDelft, Netherlands.

ABSTRACT
Anaerobic enrichment from sediments of hypersaline alkaline lakes in Wadi el Natrun (Egypt) with chitin resulted in the isolation of a fermentative haloalkaliphilic bacterium, strain ACht6-1, growing exclusively with insoluble chitin as the substrate in a sodium carbonate-based medium at pH 8.5-10.5 and total Na(+) concentrations from 0.4 to 1.75 M. The isolate had a Gram-negative cell wall and formed lipid cysts in old cultures. The chitinolytic activity was associated with cells. Analysis of the 4.4 Mb draft genome identified pathways for chitin utilization, particularly, secreted chitinases linked to the cell surface, as well as genes for the hydrolysis of other polysaccharides and fermentation of sugars, while the genes needed for aerobic and anaerobic respiration were absent. Adaptation to a haloalkaliphilic lifestyle was reflected by the gene repertoire encoding sodium rather than proton-dependent membrane-bound ion pumps, including the Rnf-type complex, oxaloacetate decarboxylase, V-type ATPase, and pyrophosphatase. The phylogenetic analysis using 16S rRNA gene and ribosomal proteins indicated that ACht6-1 forms a novel deep lineage at the class level within the bacterial candidate division TG3. Based on phylogenetic, phenotypic and genomic analyses, the novel chitinolytic bacterium is described as Chitinispirillum alkaliphilum gen. nov., sp. nov., within a novel class Chitinispirillia that could be included into the phylum Fibrobacteres.

No MeSH data available.


Related in: MedlinePlus

Cell morphology of strain ACht6-1 grown on amorphous chitin at pH 10 and 0.6 M Na+. Phase contrast microphotographs showing ACht6-1 cells bound to a chitin particle (A) and initial (B) and final (C) stages of the lipid cyst formation in free cells. Thin section electron microscopy showing ultrastructural organization of cells of strain ACht6-1 growing on chitin: actively growing cells from the exponential phase (D) and cells from the stationary phase full of PHA lipid storage (E). The image in panel C was modified from Figure 5 in Sorokin et al. (2012).
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Figure 1: Cell morphology of strain ACht6-1 grown on amorphous chitin at pH 10 and 0.6 M Na+. Phase contrast microphotographs showing ACht6-1 cells bound to a chitin particle (A) and initial (B) and final (C) stages of the lipid cyst formation in free cells. Thin section electron microscopy showing ultrastructural organization of cells of strain ACht6-1 growing on chitin: actively growing cells from the exponential phase (D) and cells from the stationary phase full of PHA lipid storage (E). The image in panel C was modified from Figure 5 in Sorokin et al. (2012).

Mentions: A composite surface sediment sample from seven hypersaline alkaline lakes in Wadi el Natrun (Libyan Desert, Egypt) was used for the isolation of anaerobic chitinolytic bacteria (Sorokin et al., 2012). The pH in the soda brines was between 9.4 and 10.1, the salinity varied between 200 and 360 g l-1 and the carbonate alkalinity varied from 0.1 to 0.75 M (Sorokin et al., 2012). Anaerobic enrichments with crystalline chitin inoculated with mixed anoxic sediments at 0.6 M of total sodium and pH 10 were dominated by two vibrio-shaped bacteria differing in size. Subculturing at 2 M total Na+ separated the small sized cells (strain ACht6-2), described previously as a member of the genus Chitinivibrio (Sorokin et al., 2014), while the larger spirillum-shaped cells grew faster at moderate salinity and were eventually isolated in pure culture by the dilution to extinction technique at 0.6 M Na+ and designated as strain ACht6-1. The isolate contained spirilloid cells motile with a single polar flagellum. In the initial growth phase, the cells were mostly attached to chitin particles. After visible chitin degradation, the cells started to massively detach from the solid phase into the liquid phase and to form round refractive bodies consisting mostly of PHB-like (Nile Blue staining) granules (Figure 1). Whilst complete chitin depolymeriszation occurred, massive cell lysis commenced, similar to that previously described for the haloalkaliphilic chitinotrophic anaerobe Chtinivibrio (Sorokin et al., 2012, 2014). It is well-documented that in Gram-negative bacteria, massive autolysis can be triggered under certain conditions, mostly in the stationary phase when the cells are losing control of their autolysins, which also include a lysozyme-like member (Holtje, 1995; Shockman et al., 1996). In our case, it might be speculated that when the substrate (chitin) is consumed, the cell-bound chitinases start to digest the cell wall murein layer of ACht6-1.


Phenotypic and Genomic Properties of Chitinispirillum alkaliphilum gen. nov., sp. nov., A Haloalkaliphilic Anaerobic Chitinolytic Bacterium Representing a Novel Class in the Phylum Fibrobacteres.

Sorokin DY, Rakitin AL, Gumerov VM, Beletsky AV, Sinninghe Damsté JS, Mardanov AV, Ravin NV - Front Microbiol (2016)

Cell morphology of strain ACht6-1 grown on amorphous chitin at pH 10 and 0.6 M Na+. Phase contrast microphotographs showing ACht6-1 cells bound to a chitin particle (A) and initial (B) and final (C) stages of the lipid cyst formation in free cells. Thin section electron microscopy showing ultrastructural organization of cells of strain ACht6-1 growing on chitin: actively growing cells from the exponential phase (D) and cells from the stationary phase full of PHA lipid storage (E). The image in panel C was modified from Figure 5 in Sorokin et al. (2012).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4814513&req=5

Figure 1: Cell morphology of strain ACht6-1 grown on amorphous chitin at pH 10 and 0.6 M Na+. Phase contrast microphotographs showing ACht6-1 cells bound to a chitin particle (A) and initial (B) and final (C) stages of the lipid cyst formation in free cells. Thin section electron microscopy showing ultrastructural organization of cells of strain ACht6-1 growing on chitin: actively growing cells from the exponential phase (D) and cells from the stationary phase full of PHA lipid storage (E). The image in panel C was modified from Figure 5 in Sorokin et al. (2012).
Mentions: A composite surface sediment sample from seven hypersaline alkaline lakes in Wadi el Natrun (Libyan Desert, Egypt) was used for the isolation of anaerobic chitinolytic bacteria (Sorokin et al., 2012). The pH in the soda brines was between 9.4 and 10.1, the salinity varied between 200 and 360 g l-1 and the carbonate alkalinity varied from 0.1 to 0.75 M (Sorokin et al., 2012). Anaerobic enrichments with crystalline chitin inoculated with mixed anoxic sediments at 0.6 M of total sodium and pH 10 were dominated by two vibrio-shaped bacteria differing in size. Subculturing at 2 M total Na+ separated the small sized cells (strain ACht6-2), described previously as a member of the genus Chitinivibrio (Sorokin et al., 2014), while the larger spirillum-shaped cells grew faster at moderate salinity and were eventually isolated in pure culture by the dilution to extinction technique at 0.6 M Na+ and designated as strain ACht6-1. The isolate contained spirilloid cells motile with a single polar flagellum. In the initial growth phase, the cells were mostly attached to chitin particles. After visible chitin degradation, the cells started to massively detach from the solid phase into the liquid phase and to form round refractive bodies consisting mostly of PHB-like (Nile Blue staining) granules (Figure 1). Whilst complete chitin depolymeriszation occurred, massive cell lysis commenced, similar to that previously described for the haloalkaliphilic chitinotrophic anaerobe Chtinivibrio (Sorokin et al., 2012, 2014). It is well-documented that in Gram-negative bacteria, massive autolysis can be triggered under certain conditions, mostly in the stationary phase when the cells are losing control of their autolysins, which also include a lysozyme-like member (Holtje, 1995; Shockman et al., 1996). In our case, it might be speculated that when the substrate (chitin) is consumed, the cell-bound chitinases start to digest the cell wall murein layer of ACht6-1.

Bottom Line: The chitinolytic activity was associated with cells.Analysis of the 4.4 Mb draft genome identified pathways for chitin utilization, particularly, secreted chitinases linked to the cell surface, as well as genes for the hydrolysis of other polysaccharides and fermentation of sugars, while the genes needed for aerobic and anaerobic respiration were absent.The phylogenetic analysis using 16S rRNA gene and ribosomal proteins indicated that ACht6-1 forms a novel deep lineage at the class level within the bacterial candidate division TG3.

View Article: PubMed Central - PubMed

Affiliation: Winogradsky Institute of Microbiology, Research Center of Biotechnology of the Russian Academy of SciencesMoscow, Russia; Department of Biotechnology, Delft University of TechnologyDelft, Netherlands.

ABSTRACT
Anaerobic enrichment from sediments of hypersaline alkaline lakes in Wadi el Natrun (Egypt) with chitin resulted in the isolation of a fermentative haloalkaliphilic bacterium, strain ACht6-1, growing exclusively with insoluble chitin as the substrate in a sodium carbonate-based medium at pH 8.5-10.5 and total Na(+) concentrations from 0.4 to 1.75 M. The isolate had a Gram-negative cell wall and formed lipid cysts in old cultures. The chitinolytic activity was associated with cells. Analysis of the 4.4 Mb draft genome identified pathways for chitin utilization, particularly, secreted chitinases linked to the cell surface, as well as genes for the hydrolysis of other polysaccharides and fermentation of sugars, while the genes needed for aerobic and anaerobic respiration were absent. Adaptation to a haloalkaliphilic lifestyle was reflected by the gene repertoire encoding sodium rather than proton-dependent membrane-bound ion pumps, including the Rnf-type complex, oxaloacetate decarboxylase, V-type ATPase, and pyrophosphatase. The phylogenetic analysis using 16S rRNA gene and ribosomal proteins indicated that ACht6-1 forms a novel deep lineage at the class level within the bacterial candidate division TG3. Based on phylogenetic, phenotypic and genomic analyses, the novel chitinolytic bacterium is described as Chitinispirillum alkaliphilum gen. nov., sp. nov., within a novel class Chitinispirillia that could be included into the phylum Fibrobacteres.

No MeSH data available.


Related in: MedlinePlus