Strategies to understand and reduce the spoilage potential of low and non-alcoholic beverages

发酵饮料被认为是“对人类消费安全”的产品，主要原因是致病微生物与最终产品无关。然而，近年来，消费者的偏好导致了“无酒精或低酒精饮料”（NABLAB’s）的流行。这一类别反映了酿造行业快速增长的市场，从2015年占所有产品的0.1%扩大到目前的5%，预计到2025年将进一步增长至33%。虽然NABLAB通常被设计为模拟酒精饮料的风味特征，但其成分却有根本的不同，并且由于营养物质浓度相对较高，pH值升高，碳酸化程度较低和乙醇减少，因此微生物污染的风险较高1。因此，NABLAB产品变质的报告急剧增加，帝亚吉欧最近的一次重大产品召回凸显了这一问题。在这个项目中，我们的目标是充分了解整个酿造过程中与NABLAB生产相关的微生物风险，包括以前被认为是低威胁的微生物污染的可能性。我们的目标是选择能够在NABLAB环境中茁壮成长的生物，并分析它们的生理和生长偏好，以通过有针对性的方法防止污染。该主题将通过如下所述的4个工作包来解决：WP 1-关键污染物的识别（0-12个月）这将通过一系列现场调查来进行，重点关注原材料、生产工艺、产品的储存和运输以及销售点的货物。为了实现这一目标，我们将利用UoN酿酒集团现有的专业知识开发一种基于宏基因组的策略（MinION），以充分捕获每个工艺阶段的微生物组。对于WP 2，还将使用标准微生物学技术分离微生物。WP 2-微生物表征（9-18个月）为了了解腐败潜力，将使用BioLog表型微阵列分析来评估NABLAB在一系列条件下的微生物生长。目的是了解微生物的生物学，包括它们对抗菌剂（啤酒花化合物，亚硫酸盐和苯甲酸盐）的适应性，它们对啤酒特性（酒精，pH值）的耐受性以及它们的营养需求（糖和必需离子）。我们已经在Biolog PM技术的应用方面拥有广泛的专业知识，以分析生长抑制剂的影响，并模拟微生物种群动态2 -3。WP 3-产品和腐败潜力之间的关系（18-27个月）在这里，我们的目标是确定所有NABLAB产品是否具有相同的腐败潜力，并确定增强或防止污染的关键属性（营养或成分）。我们将使用WP 1中开发的宏基因组方法来跟踪随时间的腐败，并通过种群监测来确定多种生物体随时间对腐败的影响。WP 4-最大限度地发挥防腐策略的影响（24-36个月）从WP 3和WP 4中获得的经验将应用于关键产品，以便使用有针对性的方法加强当前的防腐策略。其目的是减少对人工防腐剂的依赖，并提高天然特性，以防止微生物生长和腐败。

Fermented beverages are considered to be products which are 'safe for human consumption', for the primary reason that pathogenic organisms are not associated with the final product. However, in recent years, consumer preferences have led to the growth in popularity of 'no or low alcohol beverages' (NABLAB's). This category reflects a rapidly growing market within the brewing sector, expanding from <0.1% of all products in 2015 to 5% global consumption at the current time and further growth expected to reach ~33% by 2025. Although often designed to mimic alcoholic beverage flavour profiles, NABLAB's are fundamentally different in terms of their composition and are at higher risk of microbial contamination due to the relatively high concentration of nutrients, elevated pH, milder carbonation levels and reduction in ethanol1. Consequently, reports of spoilage in NABLAB products have dramatically increased, an issue highlighted by a recent major product recall by Diageo. In this project we aim to fully understand the microbial risks associated with production of NABLAB's across the brewing process, including the potential for contamination by organisms previously considered to be of low threat. We aim to select for organisms capable of thriving in the NABLAB environment and analyse their physiology and growth preferences with the ambition of preventing contamination via a targeted approach. This topic will be addressed through 4 work packages as described below: WP1 - Identification of key contaminants (0-12 months) This will be conducted via a series of site surveys focused on raw materials, the manufacturing process, storage and transport of products, and goods at point of sale. To achieve this, we will develop a metagenomic based strategy (MinION) using current expertise within the brewing group at UoN, to fully capture the microbiome at each process stage. Microbes will also be isolated using standard microbiological techniques for WP2. WP2 - Characterisation of microbes (9-18 months) To understand spoilage potential, BioLog Phenotype Microarray analysis will be used to assess microbial growth in NABLAB's under a range of conditions. The aim is to understand the biology of microorganisms, including their resilience to antimicrobials (hop compounds, sulphites and benzoates), their tolerances to beer properties (alcohol, pH) and their nutritional requirements (sugars and essential ions). We already have extensive expertise in the application of Biolog PM technology to analyse for the effect of growth inhibitors and in modelling microbial population dynamics2-3. WP3 - Relationship between product and spoilage potential (18-27 months) Here we aim to determine if all NABLAB products have the same spoilage potential and to identify key attributes (nutritional or compositional) which enhance or prevent contamination. We will use the metagenomic approach developed in WP1 to track spoilage over time and to determine the impact of multiple organisms on spoilage over time through population monitoring. WP4 - Maximising the impact of preservative strategies (24-36 months)Learnings from WP3 and WP4 will be applied to key products in order to enhance current preservation strategies using a targeted approach. The goal is to reduce dependency on artificial preservatives and to enhance natural characteristics to prevent microbial growth and spoilage.