Metaproteomic Analysis of a PAH-Degrading Dual Synthetic Bacterial Consortium Constructed with the Key Microbes of a Natural Consortium

NIETO, MACCHI, MORELLI ; MARÍA PÍA VALACCO; COPPOTELLI BM.

Online Worldwide

Congreso; World Microbe Forum brings together two of the biggest events in the microbial sciences ASM Microbe 2021 and FEMS2021, 20-24 junio 2021 Online Worldwide; 2021

ASM-ISME

Metaproteomics applied to the study of consortia is an increasingly used approach to predict roles, occurring inter-species interactions and the resulting changes in active metabolic pathways, being useful in the design of synthetic microbial cooperative consortia for biotechnological applications. A Dual Synthetic Consortium AM-Bk (DSC_AB) was constructed combining the key members (Sphingobium sp. (AM) and Burkholderia sp. (Bk)) of a natural consortium. DSC_AB exhibited higher phenanthrene (PHN) degradation efficiency than axenic cultures of AM and Bk. A previous detailed RT-qPCR analysis of PHN-related transcripts confirmed their complementation; AM would attack PHN initially and Bk would intervene in the lower pathway. The aim of this work was to apply a metaproteomic approach to unravel the interaction and regulation mechanisms that occur between each bacterium during PHN degradation. The workflow consisted of protein isolation (at 72 h) of DSC_AB, AM and Bk grown in liquid mineral medium with 200 mg / L of PHN, protein digestion and mass spectrometry (CEQUIBIEM), data acquisition and handling with XCalibur, Orbitrap, Proteome Discoverer and Perseus, searched against draft genome sequence (NHOM01: Bk, LRUK01: AM).Significantly expressed proteins (volcano plot) were identified and categorized in different metabolic pathways (BlastKoala) and complemented by manual curation. Proteins mainly categorized in xenobiotic degradation, stress and cycle cellular and transporters metabolisms underwent significant modifications in the level of expression; 9 for AM and 20 for Bk were significantly overexpressed and 39 for AM and 110 for Bk were under-expressed in DSC_AB in relation to the axenic cultures. Proteins for many steps of PHN-degradation pathway were significantly expressed in DSC-AB. Regarding their role, all under-expressed proteins of AM corresponded to the lower pathway (benzoate dioxygenase; etc.), while for Bk there was under-expression of proteins for the upper (biphenyl dioxygenase) and the lower (catechol 2,3-dioxygenase; etc.) pathways. Bk overexpressed proteins related to cell division, while AM did not. The under-expression of AM and Bk cell stress proteins (regulator PhyR, peroxidase) observed could explain the higher performance reached by the coculture, since metabolic stress is reduced in each strain. Our results confirm the cooperation between strains, which led to cellular stress reduction and increased degradation performance, yielding new criteria to combine strains in bioremediation processes.Footnotes: A mixing of the AM and Bk strains cultivated separately in the same proportion as in the DSC_AB were used as controls to discount the mixing effect in the abundance calculation. All proteins result files were submitted to PRIDE with the accession number PXD024411.