Tata Alessandra, Massaro Andrea, Miano Brunella, Petrin Sara, Antonelli Pietro, Peruzzo Arianna, Pezzuto Alessandra, Favretti Michela, Bragolusi Marco, Zacometti Carmela, Losasso Carmen, Piro Roberto
Laboratorio di Chimica Sperimentale, Istituto Zooprofilattico Sperimentale delle Venezie, Viale Fiume 78, 36100 Vicenza, Italy.
Laboratory of Microbial Ecology and Genomics, Istituto Zooprofilattico Sperimentale delle Venezie, Viale dell'Università, 35020 Legnaro, Italy.
Foods. 2024 Jun 18;13(12):1912. doi: 10.3390/foods13121912.
Raw milk cheeses harbor complex microbial communities. Some of these microorganisms are technologically essential, but undesirable microorganisms can also be present. While most of the microbial dynamics and cross-talking studies involving interaction between food-derived bacteria have been carried out on agar plates in laboratory-controlled conditions, the present study evaluated the modulation of the resident microbiota and the changes of metabolite production directly in ripening raw milk cheese inoculated with strains. Using a proxy of the pathogenic , we aimed to establish the key microbiota players and chemical signals that characterize Latteria raw milk cheese over 60 days of ripening time. The microbiota of both the control and -inoculated cheeses was analyzed using 16S rRNA targeted amplicon sequencing, while direct analysis in real time mass spectrometry (DART-HRMS) was applied to investigate the differences in the metabolic profiles of the cheeses. The diversity analysis showed the same microbial diversity trend in both the control cheese and the inoculated cheese, while the taxonomic analysis highlighted the most representative genera of bacteria in both the control and inoculated cheese: and . On the other hand, the metabolic fingerprints revealed that the complex interactions between resident microbiota and were governed by continuously changing chemical signals. Changes in the amounts of small organic acids, hydroxyl fatty acids, and antimicrobial compounds, including pyroglutamic acid, hydroxy-isocaproic acid, malic acid, phenyllactic acid, and lactic acid, were observed over time in the -inoculated cheese. In cheese that was inoculated with , was significantly correlated with the volatile compounds carboxylbenzaldheyde and cyclohexanecarboxylic acid, while was positively correlated with some volatile and flavor compounds (cyclohexanecarboxylic acid, pyroxidal acid, aminobenzoic acid, and vanillic acid). Therefore, we determined the metabolic markers that characterize a raw milk cheese inoculated with , the changes in these markers with the ripening time, and the positive correlation of flavor and volatile compounds with the resident microbiota. This multi-omics approach could suggest innovative food safety strategies based on the enhanced management of undesirable microorganisms by means of strain selection in raw matrices and the addition of specific antimicrobial metabolites to prevent the growth of undesirable microorganisms.
生乳奶酪中存在复杂的微生物群落。其中一些微生物在工艺上是必不可少的，但也可能存在不良微生物。虽然大多数涉及食品源细菌之间相互作用的微生物动态和相互作用研究都是在实验室控制条件下的琼脂平板上进行的，但本研究直接对接种了菌株的成熟生乳奶酪中的常驻微生物群的调节和代谢产物产生的变化进行了评估。我们以致病性的一种替代物为研究对象，旨在确定在60天的成熟时间内表征拉泰里亚生乳奶酪的关键微生物群落成员和化学信号。使用靶向16S rRNA的扩增子测序分析对照奶酪和接种奶酪的微生物群，同时应用实时直接分析质谱法（DART-HRMS）研究奶酪代谢谱的差异。多样性分析表明对照奶酪和接种奶酪具有相同的微生物多样性趋势，而分类学分析突出了对照奶酪和接种奶酪中最具代表性的细菌属：和。另一方面，代谢指纹图谱显示常驻微生物群和之间的复杂相互作用受不断变化的化学信号支配。在接种奶酪中，随着时间的推移，观察到小分子有机酸、羟基脂肪酸和抗菌化合物（包括焦谷氨酸、羟基异己酸、苹果酸、苯乳酸和乳酸）含量的变化。在接种的奶酪中，与挥发性化合物羧基苯甲醛和环己烷羧酸显著相关，而与一些挥发性和风味化合物（环己烷羧酸、焦氧酸、氨基苯甲酸和香草酸）呈正相关。因此我们确定了表征接种的生乳奶酪的代谢标志物、这些标志物随成熟时间的变化以及风味和挥发性化合物与常驻微生物群的正相关关系。这种多组学方法可能会提出创新的食品安全策略，即通过在原料基质中选择菌株和添加特定的抗菌代谢物来加强对不良微生物的管理，以防止不良微生物的生长。
