Molecular characterization of the aggregate formation-inducing interaction between Marinobacter adhaerens HP15 and the diatom, Thalassiosira weissflogii
Final Report Abstract
The role of heavy metal resistance in M. adhaerens HP15 during its interaction with T. weissflogii was studied by comparing wild-type and the mutant ΔczcCBA.1/2 of M. adhaerens HP15 subjected to co-culture experiments with T. weissflogii with and without zinc addition. Under zinc stress, the release of exopolymer substances by both interaction partners and the proportion of attached bacteria increased as compared to cultures without zinc. Despite similar zinc concentrations inside exopolymer particles, the wild-type of HP15 showed a higher ability to colonize aggregates than its mutant ΔczcCBA.1/2. Microscopy revealed that attached bacteria are mainly located inside exopolymer particles rather than on the diatom surface. These results demonstrated that heavy metal resistance can help diatom-associated bacteria to colonize aggregates that may be enriched in heavy metals. Subsequently, the role of flagellar motility and chemotaxis for aggregate colonization by M. adhaerens HP15 was assessed by analyzing the non-motile mutant ΔfliC and the non-chemotactic mutant ΔcheA. At low concentrations of exopolymer particles, there was no difference observable in the percentage of attached bacteria between the wild-type and both mutants. However, at high concentrations of exopolymers significantly less cells of the mutant ΔfliC were found attached as compared to the wild-type or the mutant ΔcheA. Proteomic profiling showed that attached wild-type bacteria expressed proteins involved in amino acid metabolism at a higher level than free-living bacteria. Compared to the wild-type, the mutant ΔcheA showed signs of amino acid and nutrient starvation whereas the mutant ΔfliC expressed amino acid uptake transporters at a higher level than the wild-type. The conclusion that particularly branched chain amino acids are present in TEP as a nutrient source was confirmed by amino acid quantification. Among other amino acids, leucine, isoleucine and valine were more abundant in the amino acid pool of the attached fractions as compared to the free-living fractions. Finally, mutant M. adhaerens HP15 ΔlivK was generated, which lacks the periplasmic binding protein of a branched chain amino acid uptake transporter. Expression analysis of five livK genes showed that the function of the deleted gene is compensated by the up-regulation of two other livK genes leading to no observable fitness impairment when comparing in vitro growth and amino acid uptake patterns of M. adhaerens HP15 mutant ΔlivK to the wild-type. Additionally, gene expression showed that the analyzed livK gene does not play a major role during in vitro growth of M. adhaerens HP15 wild-type. Two other livK genes were constitutively expressed at a high level and two genes took over the function of the deleted gene. Co-cultures with T. weissflogii did not reveal a difference in aggregate colonization ability between M. adhaerens HP15 wild-type and mutant ΔlivK suggesting that other livK genes may as well take over the function under in vivo conditions. These results indicate that the studied livK gene has different functions during in vivo and in vitro growth.
Publications
- Heavy metal resistance in Marinobacter adhaerens HP15 supports colonization of transparent exopolymer particles during ist interaction with diatoms. Marine Ecology Progress Series, Vol. 658. 2021, pp. 47-57.
Will V., A. Stahl, and M.S. Ullrich
(See online at https://doi.org/10.3354/meps13558)
