Cocuzzi Riccardo, Turgay Meral, Schmidt Remo S, von Ah Ueli, Bachmann Hans-Peter, Weisskopf Laure, Fröhlich-Wyder Marie-Therese
Agroscope, Bern, Switzerland.
Department of Biology, University of Fribourg, Fribourg, Switzerland.
Appl Environ Microbiol. 2025 Jul 23;91(7):e0059925. doi: 10.1128/aem.00599-25. Epub 2025 Jun 12.
() has various biotechnological applications, notably in the ripening of Swiss-type cheese, where it utilizes the two enantiomers of lactate as the main carbon source, contributing to flavor development and eye formation. Here, we genotypically characterized two strains unable to catabolize L-lactate through whole-genome sequencing and variant calling, using FAM-14222 as the reference genome, which highlighted a mutation in the gene in both strains. This gene is part of the operon, which has been previously linked to lactate utilization in other bacterial species. Subsequently, we successfully restored the strains' ability to utilize L-lactate by following an adaptive laboratory evolution approach, which involved repeated subculturing in a medium containing L-lactate as the main carbon source. Sequencing of the gene confirmed that isolates with a restored ability to utilize L-lactate had also reverted the mutation back to wild-type, supporting the involvement of the operon in L-lactate catabolism in . Moreover, the phenotype of the two L-lactate-negative strains was confirmed under cheesemaking conditions, highlighting the potential of the strains as cheese ripening cultures.IMPORTANCELactate catabolism is of paramount importance in , particularly for its industrial applications, such as Swiss-type cheese ripening. Nevertheless, the genetic background of this metabolic process is not fully understood. In our study, we developed an adaptive laboratory evolution-based approach for the elucidation of L-lactate catabolism, starting from two strains unable to utilize L-lactate. Our results delivered experimental evidence of the role of the operon in this process, as opposed to the widespread theory of L-lactate dehydrogenase-mediated oxidation. A deeper understanding of this metabolic pathway will be beneficial for a more efficient selection of industrial strains, as well as for metabolic engineering.
（）具有多种生物技术应用，特别是在瑞士型奶酪的成熟过程中，它利用乳酸的两种对映体作为主要碳源，有助于风味的形成和眼状气孔的形成。在此，我们通过全基因组测序和变异检测对两株无法分解代谢L-乳酸的菌株进行了基因分型，以FAM-14222作为参考基因组，结果表明两株菌株的该基因均发生了突变。该基因是操纵子的一部分，此前已证明其与其他细菌物种中乳酸的利用有关。随后，我们通过适应性实验室进化方法成功恢复了菌株利用L-乳酸的能力，该方法包括在以L-乳酸作为主要碳源的培养基中反复传代培养。对该基因的测序证实，具有恢复利用L-乳酸能力的分离株也已将突变恢复为野生型，这支持了操纵子参与该菌L-乳酸分解代谢的观点。此外，在奶酪制作条件下证实了这两株L-乳酸阴性菌株的表型，突出了这些菌株作为奶酪成熟培养物的潜力。重要性乳酸分解代谢在该菌中至关重要，特别是在其工业应用方面，如瑞士型奶酪的成熟过程。然而，这一代谢过程的遗传背景尚未完全了解。在我们的研究中，我们从两株无法利用L-乳酸的菌株出发，开发了一种基于适应性实验室进化的方法来阐明L-乳酸分解代谢。我们的结果提供了实验证据，证明操纵子在这一过程中的作用，这与广泛认为的L-乳酸脱氢酶介导的氧化作用的理论相反。对这一代谢途径的更深入理解将有助于更有效地选择工业菌株以及进行代谢工程。
