Cullen Kenny (weeksprout8)

Formation of exopolysaccharides (EPS) during 2,3-butanediol (2,3-BD) fermentation by Paenibacillus polymyxa increases medium viscosity, which in turn, presents considerable technical and economic challenges to 2,3-BD downstream processing. To eliminate EPS production during 2,3-BD fermentation, we used homologous recombination to disable the EPS biosynthetic pathway in P. polymyxa The levansucrase gene which encodes levansucrase, the major enzyme responsible for EPS biosynthesis in P. polymyxa, was successfully disrupted. The P. polymyxa levansucrase null mutant produced 2.5 ± 0.1 and 1.2 ± 0.2 g/L EPS on sucrose and glucose, respectively, whereas the wildtype produced 21.7 ± 2.5 and 3.1 ± 0.0 g/L EPS on the same substrates, respectively. These levels of EPS translate to 8.7- and 2.6-fold decreases in EPS formation by the levansucrase null mutant on sucrose and glucose, respectively, relative to the wildtype, with no significant reduction in 2,3-BD production. Inactivation of EPS biosynthesis led to consideracharide (EPS) production during 2,3-BD fermentation constitutes a problem during downstream processing. Specifically, EPS negatively impacts 2,3-BD separation from the fermentation broth, thereby increasing the overall cost of 2,3-BD production. The results presented in this manuscript demonstrate that inactivation of the levansucrase gene in P. polymyxa leads to diminished EPS accumulation. Additionally, a new method for EPS assay and simple protocol employing protoplasts for enhanced transformation of P. polymyxa were developed. Overall, although our study shows that levan is not the only EPS produced by P. polymyxa, it represents a significant first step towards developing cost effective 2,3-BD fermentation, devoid of EPS-associated complications during downstream processing. Copyright © 2020 American Society for Microbiology.Grassland degradation is an ecological problem worldwide. This study aims to reveal the patterns of the variations in bacterial diversity and community structure and in nitrogen cycling functional genes along a subalpine meadow degradation gradient on the Loess Plateau, China. Meadow degradation had a significant effect on the beta diversity of soil bacterial communities (P0.05) along the meadow degradation gradient. Redundancy analysis (RDA) showed that the structure of the bacterial community was strongly correlated with total nitrogen (TN), nitrate nitrogen (NO3 --N), plant Shannon diversity, plant coverage and soil bulk density (all P less then 0.05). Moreover, the abundances of N-fixation and denitrification genes of bacterial community decreased along the degradation gradient, but the abundance of nitrification genes increased along the gradient. The structure of the set of N-cycling genes present at each site was more sensitive to subalpine meadow degradation than the structure of the total bacterial cal for nitrification. These results have implications for the restoration and reconstruction of subalpine meadow ecosystem on the Loess Plateau. Copyright © 2020 American Society for Microbiology.In Lysobacter enzymogenes (Le), Le RpfB1 and Le RpfB2 were predicted to encode acyl-CoA ligases. RpfB1 is located in the Rpf gene cluster. Interestingly, we found an RpfB1 homolog (RpfB2) outside of this canonical gene cluster, and nothing is known about its functionality or mechanism. Here, we report that Le rpfB1 and Le rpfB2 can functionally replace EcFadD in the Escherichia coli fadD mutant JW1794. RpfB activates long-chain fatty acids (n-C160 and n-C180) for the corresponding fatty acyl-CoA ligase (FCL) activity in vitro, and Glu-361 plays critical roles in the catalytic mechanism of RpfB1 and RpfB2. Deletion of rpfB1 and rpfB2 resulted in significantly increased HSAF production, and overexpression of rpfB1 or rpfB2 completely suppressed HSAF production. Deletion of rpfB1 and rpfB2 resulted in increased LeDSF3 synthesis in L. enzymogenes Overall, our results showed that changes in intracellular