Sirén Kimmo, Mak Sarah Siu Tze, Melkonian Chrats, Carøe Christian, Swiegers Jan Hendrik, Molenaar Douwe, Fischer Ulrich, Gilbert M Thomas P
Institute for Viticulture and Oenology, Dienstleistungszentrum Ländlicher Raum Rheinpfalz, Neustadt an der Weinstraße, Germany.
Department of Chemistry, University of Kaiserslautern, Kaiserslautern, Germany.
Front Microbiol. 2019 Apr 9;10:697. doi: 10.3389/fmicb.2019.00697. eCollection 2019.
Although there is an extensive tradition of research into the microbes that underlie the winemaking process, much remains to be learnt. We combined the high-throughput sequencing (HTS) tools of metabarcoding and metagenomics, to characterize how microbial communities of Riesling musts sampled at four different vineyards, and their subsequent spontaneously fermented derivatives, vary. We specifically explored community variation relating to three points: (i) how microbial communities vary by vineyard; (ii) how community biodiversity changes during alcoholic fermentation; and (iii) how microbial community varies between musts that successfully complete alcoholic fermentation and those that become 'stuck' in the process. Our metabarcoding data showed a general influence of microbial composition at the vineyard level. Two of the vineyards (4 and 5) had strikingly a change in the differential abundance of . We therefore additionally performed shotgun metagenomic sequencing on a subset of the samples to provide preliminary insights into the potential relevance of this observation, and used the data to both investigate functional potential and reconstruct draft genomes (bins). At these two vineyards, we also observed an increase in non-Saccharomycetaceae fungal functions, and a decrease in bacterial functions during the early fermentation stage. The binning results yielded 11 coherent bins, with both vineyards sharing the yeast bins and . Read recruitment and functional analysis of this data revealed that during fermentation, a high abundance of might serve as a biocontrol agent against bacteria, via a putative iron depletion pathway, and this in turn could help dominate the fermentation. During alcoholic fermentation, we observed a general decrease in biodiversity in both the metabarcoding and metagenomic data. Unexpected behavior was observed in vineyard 4 according to metagenomic analyses based on reference-based read mapping. Analysis of open reading frames using these data showed an increase of functions assigned to class Actinobacteria in the end of fermentation. Therefore, we hypothesize that bacteria might sit-and-wait until activity slows down. Complementary approaches to annotation instead of relying a single database provide more coherent information true species. Lastly, our metabarcoding data enabled us to identify a relationship between stuck fermentations and abundance. Given that robust chemical analysis indicated that although the stuck samples contained residual glucose, all fructose had been consumed, we hypothesize that this was because fructophilic , rather than , dominated these fermentations. Overall, our results showcase the different ways in which metagenomic analyses can improve our understanding of the wine alcoholic fermentation process.
尽管对葡萄酒酿造过程中涉及的微生物已有广泛的研究传统，但仍有许多有待了解之处。我们结合了宏条形码和宏基因组学的高通量测序（HTS）工具，以表征在四个不同葡萄园采集的雷司令葡萄汁的微生物群落，以及它们随后自然发酵后的衍生物是如何变化的。我们特别探讨了与三个方面相关的群落变化：（i）微生物群落在不同葡萄园之间的差异；（ii）酒精发酵过程中群落生物多样性的变化；（iii）成功完成酒精发酵的葡萄汁与发酵过程中“停滞”的葡萄汁之间微生物群落的差异。我们的宏条形码数据显示了葡萄园水平上微生物组成的总体影响。其中两个葡萄园（4号和5号）在……的差异丰度上有显著变化。因此，我们还对一部分样本进行了鸟枪法宏基因组测序，以初步了解这一观察结果的潜在相关性，并利用这些数据研究功能潜力和重建基因组草图（分箱）。在这两个葡萄园中，我们还观察到在发酵早期非酿酒酵母科真菌功能增加，细菌功能减少。分箱结果产生了11个连贯的箱，两个葡萄园共享酵母箱……和……。对这些数据的读数招募和功能分析表明，在发酵过程中，高丰度的……可能通过一种假定的铁消耗途径作为细菌的生物控制剂，这反过来可能有助于……主导发酵。在酒精发酵过程中，我们在宏条形码和宏基因组数据中都观察到生物多样性普遍下降。根据基于参考读数映射的宏基因组分析，在4号葡萄园中观察到了意外的……行为。利用这些数据对开放阅读框进行分析表明，在发酵结束时，分配给放线菌纲的功能增加。因此，我们假设细菌可能会等待，直到……的活性减缓。采用互补的注释方法而不是依赖单一数据库可以提供关于真实物种的更连贯信息。最后，我们的宏条形码数据使我们能够确定发酵停滞与……丰度之间的关系。鉴于可靠的化学分析表明，尽管停滞样本中含有残留葡萄糖，但所有果糖都已被消耗，我们假设这是因为嗜果糖的……而不是……主导了这些发酵。总体而言，我们的结果展示了宏基因组分析可以增进我们对葡萄酒酒精发酵过程理解的不同方式。
