Food Bioscience Department, Teagasc Food Research Centre, Cork, Ireland.
School of Microbiology, University College Cork, Cork, Ireland.
Appl Environ Microbiol. 2018 Oct 1;84(20). doi: 10.1128/AEM.01305-18. Print 2018 Oct 15.
Spoilage and pathogenic spore-forming bacteria are a major cause of concern for producers of dairy products. Traditional agar-based detection methods employed by the dairy industry have limitations with respect to their sensitivity and specificity. The aim of this study was to identify low-abundance sporeformers in samples of a powdered dairy product, whey powder, produced monthly over 1 year, using novel culture-independent shotgun metagenomics-based approaches. Although mesophilic sporeformers were the main target of this study, in one instance thermophilic sporeformers were also targeted using this culture-independent approach. For comparative purposes, mesophilic and thermophilic sporeformers were also tested for within the same sample using culture-based approaches. Ultimately, the approaches taken highlighted differences in the taxa identified due to treatment and isolation methods. Despite this, low levels of transient, mesophilic, and in some cases potentially pathogenic sporeformers were consistently detected in powder samples. Although the specific sporeformers changed from one month to the next, it was apparent that 3 groups of mesophilic sporeformers, namely, , /, and a third, more heterogeneous group containing , dominated across the 12 samples. Total thermophilic sporeformer taxonomy was considerably different from mesophilic taxonomy, as well as from the culturable thermophilic taxonomy, in the one sample analyzed by all four approaches. Ultimately, through the application of shotgun metagenomic sequencing to dairy powders, the potential for this technology to facilitate the detection of undesirable bacteria present in these food ingredients is highlighted. The ability of sporeformers to remain dormant in a desiccated state is of concern from a safety and spoilage perspective in dairy powder. Traditional culturing techniques are slow and provide little information without further investigation. We describe the identification of mesophilic sporeformers present in powders produced over 1 year, using novel shotgun metagenomic sequencing. This method allows detection and identification of possible pathogens and spoilage bacteria in parallel. Strain-level analysis and functional gene analysis, such as identification of toxin genes, were also performed. This approach has the potential to be of great value with respect to the detection of spore-forming bacteria and could allow a processor to make an informed decision surrounding process changes to reduce the risk of spore contamination.
变质和致病孢子形成细菌是乳制品生产者关注的主要问题。乳制品行业使用的传统基于琼脂的检测方法在灵敏度和特异性方面存在局限性。本研究的目的是使用新的非培养基于 shotgun 宏基因组学的方法来鉴定粉状乳制品（乳清粉）样本中低丰度的孢子形成菌。虽然嗜温孢子形成菌是本研究的主要目标，但在一种情况下，也使用这种非培养方法靶向耐热孢子形成菌。出于比较目的，还使用基于培养的方法在同一样品中测试了嗜温和耐热孢子形成菌。最终，所采用的方法突出了由于处理和分离方法而导致的分类群差异。尽管如此，在粉末样品中仍持续检测到低水平的瞬态、嗜温且在某些情况下可能是致病的孢子形成菌。尽管特定的孢子形成菌逐月变化，但显然有 3 组嗜温孢子形成菌，即 / 、 / 和第 3 组更具异质性的组，其中包含 、 ，在 12 个样品中占主导地位。通过所有 4 种方法分析的一个样品中的总耐热孢子形成菌分类与嗜温分类以及可培养耐热分类有很大不同。最终，通过将 shotgun 宏基因组测序应用于乳制品粉末，突出了该技术在检测这些食品成分中存在的不良细菌方面的潜力。从乳制品粉末的安全性和变质角度来看，孢子形成菌在干燥状态下保持休眠的能力令人担忧。传统的培养技术速度较慢，并且在没有进一步调查的情况下提供的信息很少。我们描述了使用新型 shotgun 宏基因组测序鉴定在 1 年以上生产的粉末中存在的嗜温孢子形成菌。这种方法允许同时检测和鉴定可能的病原体和腐败细菌。还进行了菌株水平分析和功能基因分析，例如鉴定毒素基因。这种方法对于检测孢子形成菌具有巨大的价值，并可以使加工商能够就减少孢子污染风险的工艺变更做出明智的决策。
