Halanaerobium polyolivorans sp. nov.-A Novel Halophilic Alkalitolerant Bacterium Capable of Polyol Degradation: Physiological Properties and Genomic Insights

Abstract

A search for the microorganisms responsible for the anaerobic degradation of osmoprotectants in soda lakes resulted in the isolation of a novel halophilic and alkalitolerant strain, designated Z-7514^T. The cells were Gram-stain-negative and non-endospore-forming rods. Optimal growth occurs at 1.6-2.1 M Na⁺, pH 8.0-8.5, and 31-35 °C. The strain utilized mainly sugars, low molecular polyols, and ethanolamine as well. The G+C content of the genomic DNA of strain Z-7514^T was 33.3 mol%. Phylogenetic and phylogenomic analyses revealed that strain Z-7514^T belongs to the genus Halanaerobium. On the basis of phenotypic properties and the dDDH and ANI values with close validly published species, it was proposed to evolve strain Z-7514^T within the genus Halanaerobium into novel species, for which the name Halanaerobium polyolivorans sp. nov. was proposed. The type strain was Z-7514^T (=KCTC 25405^T = VKM B-3577^T). For species of the genus Halanaerobium, the utilization of ethylene glycol, propylene glycol, and ethanolamine were shown for the first time. The anaerobic degradation of glycols and ethanolamine by strain Z-7514^T may represent a novel metabiotic pathway within the alkaliphilic microbial community. Based on a detailed genomic analysis, the main pathways of catabolism of most of the used substrates have been identified.

Keywords: Halanaerobium; alkaliphilic microbial community; ethanolamine; glycerol; polyol degradation.

Figure 1

View of strain Z-7514^T grown on glycerol: (A) phase-contrast light microscope image. Scale bar, 10 µm; (B) electron micrograph image. Arrows point to bacterial microcompartments (BMC). Scale bar, 1 µm.

Figure 2

Maximum-likelihood phylogenetic tree based on 16S rRNA gene sequences (1435 nucleotide sites) reconstructed with evolutionary model GTR+I+G4+F, showing the position of strain Z-7514^T with closely related members of the genus Halanaerobium. Bootstrap values (>50%) are listed as percentages at the branching points. GenBank accession numbers for 16S rRNA genes are indicated in brackets. Bar, 0.05 substitutions per nucleotide position.

Figure 3

Maximum-likelihood phylogenomic tree derived from concatenated 120 single copy marker proteins showing the position of strain Z-7514^T in relation to taxonomically characterized members of the order Halanaerobiales. Phylogenomic analysis was performed with an LG+F+I+G4 model based on 34747 amino acid positions. The tree was rooted using Bacillus subtilis NCIB 3610^T as the outgroup. Accession numbers for the genomes are indicated in brackets. Bar, 0.1 amino acid substitutions per site.

Figure 4

Number of genes associated with the general COG functional categories.

Figure 5

Genetic organization of the pdu (A) and eut (B) gene clusters in strain Z-7514^T and Salmonella enterica subsp. Enterica LT2. Colors: red—a pin protein used as a query in the GizmoGene server; white—hypothetical protein; the others—the same color in sequences identifies the protein members of the same family. The functions of some encoded proteins are indicated. The letters between gene sequences refer to pdu and eut genes, i.e A, B, C, etc., means pduA, pduB, pduC, etc. (panel (A)) and eutA, eutB, eutC, etc. (panel (B)), respectively. Designations outside sequences are: BMC, bacterial microcompartment; ADH, alcohol dehydrogenase; ACK, acetate kinase; PDF, propanediol diffusion facilitator.

Figure 6

A tentative scheme of substrates’ utilization by strain Z-7514^T. Substrates and products are marked in bold. Intermediates are marked in italic. Abbreviations for substances: EA—ethanolamine, EG—ethylene glycol, 1,2-PD—propylene glycol, 3-HPA—3-hydroxypropanal, 1,3-PD—1,3-propanediol. Genes and enzymes: pduCDE—1,2-propanediol dehydratase (EC 4.2.1.28), gldA—glycerol dehydrogenase (EC 1.1.1.6), glpK—glycerol kinase (EC 2.7.1.30), gutB—L-iditol (D-sorbitol) 2-dehydrogenase (EC 1.1.1.14), eutBC—ethanolamine ammonia lyase (EC 4.3.1.7), tktA—transketolase (EC 2.2.1.1), pduP—propionaldehyde dehydrogenase (EC 1.2.1.87), dhaT—1,3-propanediol dehydrogenase (EC 1.1.1.202), ldh—lactate dehydrogenase (EC 1.1.1.27), nifJ—pyruvate/ferredoxin oxidoreductase (EC:1.2.7.1), pta—phosphate acetyltransferase (EC 2.3.1.8), ackA—acetate kinase (EC 2.7.2.1), fakAB—nonspecific fatty acid kinase (EC 2.7.2.18). *—NAD⁺ dependent reaction.