title: Flocculation-Related Gene Identification by Whole-Genome Sequencing of Thauera aminoaromatica MZ1T Floc-Defective Mutants.
abstract: Thauera aminoaromatica MZ1T, a floc-forming bacterium isolated from an industrial activated-sludge wastewater treatment plant, overproduces exopolysaccharide (EPS), leading to viscous bulking. This phenomenon results in poor sludge settling and dewatering during the clarification process. To identify genes responsible for bacterial flocculation, a whole-genome phenotypic-sequencing technique was applied. Genomic DNA of MZ1T flocculation-deficient mutants was subjected to massively parallel sequencing. The resultant high-quality reads were assembled and compared to the reference genome of the wild type (WT). We identified nine nonsynonymous mutations and one nonsense mutation putatively involved in EPS biosynthesis. Complementation of the nonsense mutation located in an EPS deacetylase gene restored the flocculating phenotype. The Fourier transform infrared (FTIR) spectra of EPS isolated from the wild type showed a reduced C=O peak of the N-acetyl group at 1,665 cm(-1) compared to the spectra of MZ1T floc-deficient mutant EPS, suggesting that the WT EPS was partially deacetylated. Gene expression analysis also demonstrated that the putative deacetylase gene transcript increased before flocculation occurred. These data suggest that targeting deacetylation processes via direct chemical modification of EPS or enzyme inhibition may prove useful in combating viscous bulking in this and related bacteria.
authors.affiliation: Department of Biological Sciences, University of North Texas, Denton, Texas, USA Michael.Allen@unthsc.edu.
authors.forename: P
authors.initials: P
authors.lastname: Prombutara
authors.affiliation: Department of Biological Sciences, University of North Texas, Denton, Texas, USA Center for Biosafety and Biosecurity, University of North Texas Health Science Center, Fort Worth, Texas, USA Department of Molecular and Medical Genetics, University of North Texas Health Science Center, Fort Worth, Texas, USA.
authors.forename: M S
authors.initials: MS
authors.lastname: Allen
mesh_terms: D018829:Adhesins, Bacterial / Q000235:genetics* / Q000378:metabolism; D000581:Amidohydrolases / Q000235:genetics / Q000378:metabolism; D001422:Bacterial Adhesion; D018389:Codon, Nonsense; D005816:Genetic Complementation Test; D016680:Genome, Bacterial; D059014:High-Throughput Nucleotide Sequencing; D017354:Point Mutation; D011135:Polysaccharides, Bacterial / Q000737:chemistry / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017550:Spectroscopy, Fourier Transform Infrared; D020628:Thauera / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism / Q000502:physiology
chemical_list: D018829:Adhesins, Bacterial; D018389:Codon, Nonsense; D011135:Polysaccharides, Bacterial; D000581:Amidohydrolases; C022427:polysaccharide deacetylase