3-Methylbutanal is a key volatile compound that imparts a nutty flavor to Cheddar cheese. Lactococcus lactis has been successfully applied as a starter to increase the level of 3-methylbutanal produced during the ripening of cheese. However, the mechanism of action and genetic diversity of this bacterium for 3-methylbutanal biosynthesis remains unclear. In this study, we investigated the association between the L. lactis genotype and phenotype in the biosynthesis of 3-methylbutanal via both direct and indirect pathways. Fourteen strains of L. lactis were screened for the capacity to produce 3-methylbutanal, and strain 408 (>140 μM) produced the highest among all tested strains, which exhibited both α-keto acid decarboxylase and α-ketoacid dehydrogenase activities. Furthermore, the results of a sodium meta-arsenite inhibition experiment showed that the 3-methylbutanal–producing capacities of each strain declined to various degrees. The kdcA gene, which encodes the direct pathway component α-ketoacid decarboxylase, was detected in 4 of the 14 strains, of which only strain 408 contained the full-length gene. We then characterized the genes associated with the indirect pathway by detecting the expression levels of the pdh gene cluster, ack, and pta, which were expressed at relatively higher levels in a high-yield strain than in a low-yield strain. As a result, these L. lactis strains were divided into 3 categories according to gene diversity, gene expression, and 3-methylbutanal production. The results of this study refine our knowledge of the genetic determinants of 3-methylbutanal biosynthesis in L. lactis and explain the effect of both synthesis pathways on 3-methylbutanal production.

3-甲基丁醛是一种关键的挥发性化合物，可为切达干酪赋予坚果风味。乳酸乳球菌已成功用作发酵剂，以提高奶酪成熟过程中产生的 3-甲基丁醛水平。然而，这种细菌对 3-甲基丁醛生物合成的作用机制和遗传多样性仍不清楚。在这项研究中，我们通过直接和间接途径研究了 3-甲基丁醛生物合成中L. lactis基因型和表型之间的关联。筛选了14 株L.lactis产生 3-甲基丁醛的能力，菌株 408 (>140 μ M) 在所有测试菌株中产量最高，同时表现出 α-酮酸脱羧酶和 α-酮酸脱氢酶活性。此外，偏亚砷酸钠抑制实验的结果表明，每个菌株的 3-甲基丁醛生产能力都有不同程度的下降。的KDCA基因，其编码的直接途径组分α酮酸脱羧酶，在14株，其中只有菌株408包含的全长基因的4进行检测。然后，我们通过检测pdh基因簇、ack和pta的表达水平来表征与间接途径相关的基因。，其在高产菌株中的表达水平高于低产菌株。结果，这些乳酸乳球菌菌株根据基因多样性、基因表达和 3-甲基丁醛产量分为 3 类。本研究的结果完善了我们对乳酸乳球菌3-甲基丁醛生物合成的遗传决定因素的了解，并解释了两种合成途径对 3-甲基丁醛生产的影响。