Functional characterization of ligninolytic Klebsiella spp. strains associated with soil and freshwater

Abstract

Overcoming recalcitrance of lignin has motivated bioprospecting of high-yielding enzymes from environmental ligninolytic microorganisms associated with lignocellulose degrading-systems. Here, we performed isolation of 21 ligninolytic strains belonging to the genus Klebsiella spp., driven by the presence of lignin in the media. The fastest-growing strains (FP10-5.23, FP10-5.22 and P3TM1) reached the stationary phase in approximately 24 h, in the media containing lignin as the main carbon source. The strains showed biochemical evidence of ligninolytic potential in liquid- and solid media-converting dyes, which the molecular structures are similar to lignin fragments. In liquid medium, higher levels of dye decolorization was observed for P3TM.1 in the presence of methylene blue, reaching 98% decolorization in 48 h. The highest index values (1.25) were found for isolates P3TM.1 and FP10-5.23, in the presence of toluidine blue. The genomic analysis revealed the presence of more than 20 genes associated with known prokaryotic lignin-degrading systems. Identification of peroxidases (lignin peroxidase—LiP, dye-decolorizing peroxidase—DyP, manganese peroxidase—MnP) and auxiliary activities (AA2, AA3, AA6 and AA10 families) among the genetic repertoire suggest the ability to produce extracellular enzymes able to attack phenolic and non-phenolic lignin structures. Our results suggest that the Klebsiella spp. associated with fresh water and soil may play important role in the cycling of recalcitrant molecules in the Caatinga (desert-like Brazilian biome), and represent a potential source of lignin-degrading enzymes with biotechnological applications.

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Acknowledgements

This work was supported by the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) (475088/2012-3-APQ). Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) (No. 23038.009420/2012-71), Fundação de Amparo à Pesquisa do Estado da Bahia (FAPESB) (JCB0064/2013) and National Science Foundation (13330800).

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Correspondence to Thiago Bruce.

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Communicated by Erko Stackebrandt.

Electronic supplementary material

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Supplementary Table 1: Similarity between 16S rRNA gene sequences of Klebsiella isolates relative to the nearest phylogenetic neighbors. Strains isolated from soil samples have names beginning with the letter “P”, and strains isolated from freshwater samples have names beginning with the letter “F” (PNG 28 KB)

Supplementary Figure S1: Growth curve of isolates FP10-5.22 (Klebsiella variicola), FP10-5.23 (K. oxytoca), and P3TM.1 (K. variicola) in medium containing lignin as the sole carbon source (PNG 42 KB)

Supplementary Table 2: Functions associated with lignin degradation found in the Klebsiella spp. genomes, based on RAST annotation (PNG 580 KB)

Supplementary Figure S2: Catalase test for FP10-5.22 (Klebsiella variicola), FP10-5.23 (K. oxytoca), and P3TM.1 (K. variicola). B represents the blank, without presence of microbial culture (JPG 3567 KB)

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Melo-Nascimento, A.O.d.S., Treumann, C., Neves, C. et al. Functional characterization of ligninolytic Klebsiella spp. strains associated with soil and freshwater. Arch Microbiol 200, 1267–1278 (2018). https://doi.org/10.1007/s00203-018-1532-0

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Keywords

  • Lignocellulosic biomass
  • Lignin
  • Klebsiella
  • Biodegradation
  • Bioprospecting
  • Lignin-degrading enzymes