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Baby chicks administered a fecal transplant from adult chickens are resistant to Salmonella colonization by competitive exclusion. A two-pronged approach was used to investigate the mechanism of this process. First, Salmonella response to an exclusive (Salmonella competitive exclusion product, Aviguard®) or permissive microbial community (chicken cecal contents from colonized birds containing 7.85 Log10 Salmonella genomes/gram) was assessed ex vivo using a S. typhimurium reporter strain with fluorescent YFP and CFP gene fusions to rrn and hilA operon, respectively. Second, cecal transcriptome analysis was used to assess the cecal communities' response to Salmonella in chickens with low (≤5.85 Log10 genomes/g) or high (≥6.00 Log10 genomes/g) Salmonella colonization. The ex vivo experiment revealed a reduction in Salmonella growth and hilA expression following co-culture with the exclusive community. The exclusive community also repressed Salmonella's SPI-1 virulence genes and LPS modification, while the anti-virulence/inflammatory gene avrA was upregulated. Salmonella transcriptome analysis revealed significant metabolic disparities in Salmonella grown with the two different communities. Propanediol utilization and vitamin B12 synthesis were central to Salmonella metabolism co-cultured with either community, and mutations in propanediol and vitamin B12 metabolism altered Salmonella growth in the exclusive community. There were significant differences in the cecal community's stress response to Salmonella colonization. Cecal community transcripts indicated that antimicrobials were central to the type of stress response detected in the low Salmonella abundance community, suggesting antagonism involved in Salmonella exclusion. This study indicates complex community interactions that modulate Salmonella metabolism and pathogenic behavior and reduce growth through antagonism may be key to exclusion. [ABSTRACT FROM AUTHOR]

Reversible lysine acylation (RLA) is a conserved posttranslational modification that cells of all domains of life use to regulate the biological function of proteins, some of which have enzymatic activity. Many AMP-forming organic acid:CoA ligases are regulated via acylation in prokaryotes and eukaryotes. Here, we report the acetylation of the o -succinylbenzoyl-CoA synthetase (EC 6.2.1.26) of Bacillus subtilis ( Bs MenE) by the GCN5-related acetyltransferase (GNAT) AcuA enzyme of this bacterium. Bs MenE is part of the metabolic pathway that assembles menaquinone (MK), an essential component of the electron transport chain in B. subtilis . We demonstrate that the active-site lysine 471 (K471) of Bs MenE is acetylated specifically by Bs AcuA, and that acetylated Bs MenE ( Bs MenE Ac ) is deacetylated by the NAD + -dependent sirtuin ( Bs SrtN) of this bacterium. The in vivo analyses performed in this study were done in an Escherichia coli Δ menE strain because the enzymatic activity of MenE is essential in B. subtilis , but not in E. coli . The use of a heterologous system allowed us to assess the effect of acetylation on Bs MenE function under MK-dependent growth conditions. Based on our in vivo data, we suggest that regulation of Bs MenE by RLA reduces MK production, negatively affecting the growth rate and yield of the culture.IMPORTANCEReversible lysine acylation (RLA) is a posttranslational modification used by all cells to rapidly control the biological function of proteins. Herein, we identify an acetyltransferase and deacetylase in the soil bacterium Bacillus subtilis that can modify/demodify an enzyme required for the synthesis of menaquinone (MK), an essential electron carrier involved in respiration in cells of all domains of life. Based on our data, we suggest that under some as-yet-undefined physiological conditions, B. subtilis modulates MK biosynthesis, which changes the flux of electrons through the electron transport chain of this bacterium. To our knowledge, this is the first example of control of respiration by RLA.

Cobamides (Cbas) are cobalt-containing cyclic tetrapyrroles used by cells from all domains of life as co-catalyst of diverse reactions. There are several structural features that distinguish Cbas from one another. The most relevant of those features discussed in this review is the lower ligand, which is the nucleobase of a ribotide located in the lower face of the cyclic tetrapyrrole ring. The above-mentioned ribotide is known as the nucleotide loop, which is attached to the ring by a short linker. In Cbas, the nucleobase of the ribotide can be benzimidazole or derivatives of it, purine or derivatives of it, or phenolic compounds. Given the importance of Cbas in prokaryotic metabolism, it is not surprising that prokaryotes have evolved enzymes that cleave part or the entire nucleotide loop. This function is advantageous when Cbas contain nucleobases that somehow interfere with the function of Cba-dependent enzymes in the organism. After cleavage, Cbas are rebuilt via the nucleotide loop assembly (NLA) pathway, which includes enzymes that activate the nucleobase and the ring intermediate, followed by condensation of activated intermediates and a final dephosphorylation reaction. This exchange of nucleobases is known as Cba remodeling. The NLA pathway is used to salvage Cba precursors from the environment.