Artificial consortia of Bacillus amyloliquefaciens HM618 and Bacillus subtilis for utilizing food waste to synthetize iturin A.

Food waste is a cheap and abundant organic resource that can be used as a substrate for the production of the broad-spectrum antifungal compound iturin A. To increase the efficiency of food waste biotransformation, different artificial consortia incorporating the iturin A producer Bacillus amyloliquefaciens HM618 together with engineered Bacillus subtilis WB800N producing lipase or amylase were constructed. The results showed that recombinant B. subtilis WB-A13 had the highest amylase activity of 23406.4 U/mL, and that the lipase activity of recombinant B. subtilis WB-L01 was 57.5 U/mL. When strain HM618 was co-cultured with strain WB-A14, the higher yield of iturin A reached to 7.66 mg/L, representing a 32.9% increase compared to the pure culture of strain HM618. In the three-strain consortium comprising strains HM618, WB-L02, and WB-A14 with initial OD600 values of 0.2, 0.15, and 0.15, respectively, the yield of iturin A reached 8.12 mg/L, which was 38.6% higher than the control. Taken together, artificial consortia of B. amyloliquefaciens and recombinant B. subtilis can produce an increased yield of iturin A, which provides a new strategy for the valorization of food waste.

Co-culture of Bacillus amyloliquefaciens and recombinant Pichia pastoris for utilizing kitchen waste to produce fengycins.

Kitchen waste (KW) is a vast potential source of fermentable substrates. To bio-convert the KW into high-value chemicals, we used KW as substrate for the production of fengycin by an artificial consortium containing Bacillus amyloliquefaciens HM618 producing fengycin and the engineering Pichia pastoris producing amylase, glucosidase, or lipases. The maximal amylase activity of the constructed amylase-producing engineering strain (recombinant P. pastoris GS115-amy98) reached 385.4 U‧mL-1. The engineering strain GS115-α-glu53 producing glucosidase reached an enzyme activity titer of 247.3 U‧mL-1, while the lipase activities of the engineering strains GS115-lip2, GS115-α-lip2, and GS115-lip7 were around 90.0 U‧mL-1, with no significant differences among them. Liquid chromatography-mass spectrometry (LC-MS) analysis showed that the components of fengycin synthesized by B. amyloliquefaciens HM618 were complex, including C14-C18 fengycins A, C13-C14 fengycins B, C16-C18 fengycins B, C16 fengycin B2 and some fengycin homologues with unsaturated fatty acid chains. The levels of fengycin were 15.9 mg‧L-1 and 4.6 mg‧L-1 under the co-culture with strain HM618 and the recombinant strains producing amylase and lipase, respectively. The maximal titer of fengycin was 21.2 mg‧L-1 in the artificial consortia consisting of HM618 and the engineering strains producing glucosidase, amylase and lipase. Taken together, these results show that the co-culture of B. amyloliquefaciens HM618 and engineering strains producing amylase and lipase can promote the conversion of KW into fengycin. The work provides a new strategy for boosting the resource utilization of KW.

Improved the lipopeptide production of Bacillus amyloliquefaciens HM618 under co-culture with the recombinant Corynebacterium glutamicum producing high-level proline.

The application of antibacterial lipopeptides is limited by high cost and low yield. Herein, the exogenous L-proline significantly improved lipopeptide production by Bacillus amyloliquefaciens HM618. A recombinant Corynebacterium glutamicum producing high levels of proline using genetically modifying proB and putA was used to establish consortium, to improve lipopeptide production of strain HM618. Compared to a pure culture, the levels of iturin A, fengycin, and surfactin in consortium reached 67.75, 39.32, and 37.25 mg L-1, respectively, an increase of 3.19-, 2.05-, and 1.63-fold over that produced by co-cultures of B. amyloliquefaciens and recombinant C. glutamicum with normal medium. Commercial amylase and recombinant Pichia pastoris with a heterologous amylase gene were used to hydrolyze kitchen waste. A three-strain consortium with recombinant P. pastoris and C. glutamicum increased the lipopeptide production of strain HM618 in medium containing KW. This work provides new strategies to improve lipopeptide production by B. amyloliquefaciens.