ABSTRACT The production of gueuze beers through refermentation and maturation of blends of lambic beer in bottles is a way for lambic brewers to cope with the variability among different lambic beer batches. The resulting gueuze beers are more carbonated than lambic beers and are supposed to possess a unique flavor profile that varies over time. To map this refermentation and maturation process for gueuze production, a blend of lambic beers was made and bottled, whereby one of them was produced with the old wheat landrace Zeeuwse Witte . Through the use of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and high-throughput sequencing of bacterial and fungal amplicons, in combination with metabolite target analysis, new insights into gueuze production were obtained. During the initial stages of refermentation, the conditions in the bottles were similar to those encountered during the maturation phase of lambic beer productions in wooden barrels, which was also reflected microbiologically (presence of Brettanomyces species, Pediococcus damnosus , and Acetobacter lambici ) and biochemically (ethanol, higher alcohols, lactic acid, acetic acid, volatile phenolic compounds, and ethyl esters). However, after a few weeks of maturation, a switch from a favorable environment to one with nutrient and dissolved oxygen depletion resulted in several changes. Concerning the microbiology, a sequential prevalence of three lactic acid bacterial species occurred, namely, P. damnosus , Lentilactobacillus buchneri , and Lactobacillus acetotolerans , while the diversity of the yeasts decreased. Concerning the metabolites produced, mainly those of the Brettanomyces yeasts determined the metabolic profiles encountered during later stages of the gueuze production. IMPORTANCE Gueuze beers are the result of a refermentation and maturation process of a blend of lambic beers carried out in bottles. These gueuze beers are known to have a long shelf life, and their quality typically varies over time. However, knowledge about gueuze production in bottles is scarce. The present study provided more insights into the varying microbial and metabolite composition of gueuze beers during the first 2 years of this refermentation and maturation process. This will allow gueuze producers to gain more information about the influence of the refermentation and maturation time on their beers. These insights can also be used by gueuze producers to better inform their customers about the quality of young and old gueuze beers.

Belgium is known for its traditional lambic beer productions, obtained through spontaneous fermentation and maturation in wooden barrels. Lambic beer is also used to make fruit lambic beers, such as Kriek beer. Despite fruit beer being an old beer type, dating back to the second half of the seventeenth century, no research has been performed on lambic beer-fruit co-fermentation processes. Further, these beers get competition from market-driven, sweet, (fruit-)flavored ones without the co-fermentation step. This paper will first discuss a new, general fruit beer classification, going from sour fruit beers produced through co-fermentation to sweet ones without a co-fermentation step. Second, a state-of-the-art of the scarce literature on the microbiology and metabolomics of lambic beer-fruit co-fermentation processes will be given.

Polyphasic taxonomic and comparative genomic analyses revealed that a series of lambic beer isolates including strain LMG 32668T and the kombucha isolate LMG 32879 represent a novel species among the acetic acid bacteria, with Acidomonas methanolica as the nearest phylogenomic neighbor with a valid name. Overall genomic relatedness indices and phylogenomic and physiological analyses revealed that this novel species was best classified in a novel genus for which we propose the name Brytella acorum gen. nov., sp. nov., with LMG 32668T (=CECT 30723T) as the type strain. The B. acorum genomes encode a complete but modified tricarboxylic acid cycle, and complete pentose phosphate, pyruvate oxidation and gluconeogenesis pathways. The absence of 6-phosphofructokinase which rendered the glycolysis pathway non-functional, and an energy metabolism that included both aerobic respiration and oxidative fermentation are typical metabolic characteristics of acetic acid bacteria. Neither genome encodes nitrogen fixation or nitrate reduction genes, but both genomes encode genes for the biosynthesis of a broad range of amino acids. Antibiotic resistance genes or virulence factors are absent.

Lambic beers are beers produced through spontaneous fermentation and maturation in wooden barrels. The production process of lambic beers differs from the production processes of lagers and ales in process technology, environmental parameters, and the use of specific raw materials. Moreover, every lambic beer production process is unique in terms of microbiology and flavor formation because of its dependence on the spontaneous inoculation of microorganisms coming from the environmental air (contacting the open coolship and other brewery equipment) and the inner surfaces of the barrels. Several factors influence the inter- and intraspecies microbial successions during lambic beer wort fermentation and maturation and determine the final quality of the end products. The possibility to manually acidify the wort, the presence of species-specific metabolic traits, the environmental temperature, the co-occurrence of lactic acid bacteria and acetic acid bacteria, as well as yeasts, and the quality of the wooden barrels all determine the progress and outcome of the lambic beer production process. Further alterations in quality and flavor of lambic beers can be achieved by blending practices and additional bottle refermentations. This results in a vast array of lambic-derived beer products (e.g., gueuze) with complex taste and aroma profiles and specific characteristics, which separate them from most other commercially available beers.