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While most beer styles are fermented using one culture of ''S. Saccharomyces cerevisiae'' or ''S. pastorianus'', Flanders Red Ales are fermented with a [[Mixed Fermentation|mixed culture fermentation]] and sometimes with a [[Spontaneous Fermentation|spontaneous fermentation]] . At one brewery studied by Martens et al., two beers were produced using mixed fermentation and blended together. The first "light beer" was 11°P and was less acidic, while the second "heavy beer" was 13°P and served as an Old Ale if unblended. Both beers were inoculated with an acid washed yeast slurry that was harvested from a previous fermentation of the "light beer", and contained about 5% lactic acid bacteria. The fermentation of these beers followed three stages: 1. a seven day ethanol fermentation that is dominated by ''Saccharomyces'', 2. a four to five week lactic acid fermentation that was dominated by ''Lactobacilli'', and 3. a twenty to twenty-four month fermentation that was dominated by ''Brettanomyces'', ''Lactobacilli'', ''Pediococcus parvulus'', and acetic acid bacteria. The development of the ''Brettanomyces'' and ''Pediococcus'' stage was similar to the development of these microbes in [[Lambic]] fermentation. The "light beer" was never allowed to go through the third phase of fermentation, and was instead chilled to 0°C and then used to blend with previous batches of the "heavy beer" <ref name="Martens">[http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1997.tb00939.x/abstract MICROBIOLOGICAL ASPECTS OF A MIXED YEAST—BACTERIAL FERMENTATION IN THE PRODUCTION OF A SPECIAL BELGIAN ACIDIC ALE. H. Martens, D. Iserentant andH. Verachtert. 1997.]</ref>. Interestingly, and perhaps frustratingly, Flanders red and brown ales have been the subject of published studies far less than Belgian lambic beers, so the following information is based off of the Martens et al. study (see reference).  ====Primary Fermentation====In the brewery studied by Martens et al., the "light beer" was inoculated with a harvested yeast slurry of multiple strains of ''S. cerevisiae'' at a rate of 8x10⁶ CFU/mL, andthe "heavy beer" was inoculated with 1x10⁷ CFU/mL. Small numbers of ''Candida guillermondii'' and ''Candida datilla'' were reportedly in the yeast slurry, but their identification was questioned in the study and they were not found during primary fermentation. One interesting finding was that the ''S. cerevisiae'' strains used at this brewery formed sexual spores ([https://en.wikipedia.org/wiki/Ascospore ascospore]), which is quite unusual for brewing yeasts. The yeast in the "heavy beer" grew slower (3 days) and reached an overall cell count that was lower than the "light beer", which reached maximum cell count in 2 days. Yeast slurries with more lactic acid bacteria are generally used to inoculate the "heavy beer", and this may retard the yeast growth in the "heavy beer". The harvested slurry is always taken from the "light beer", which may be less adapted to the fermentation of the "heavy beer". Also, after 1 week the yeast flocculated and settled out better in the "light beer" than they did in the "heavy beer". Although lactic acid bacteria were in the yeast slurry, their growth only started to appear after 4 days into the primary fermentation. No enterobacteria or acetic acid bacteria were found during this first phase of fermentation <ref name="Martens"></ref>.  The lactic acid bacteria found in yeast slurry were 18 strains of ''L. delbruekii ssp delbruekii'' and 12 strains of ''L. delbruekii ssp bulgaricus''. Small amounts of ''Pediococcus'' were also identified in the slurry, but were impossible to isolate with enrichment until they were found in the primary fermentation. During primary fermentation, one strain of ''L. platnarum'', two strains of ''L. brevis'', and one strain of ''Pediococcus parvulus'' were identified <ref name="Martens"></ref>.  ====Secondary Fermentation====After the 7 days of the primary fermentation, the beer was transferred to a secondary fermenter. Both the yeast and bacteria populations saw a decline during the transfer, and then a small and gradual growth in secondary with the final yeast count being 4.3x10⁵ in the "heavy beer" and 6.4x10³ in the "light beer". Lactic acid bacteria grew much faster and became dominate in the "light beer", whereas in the "heavy beer" they grew more slowly and yeast remained the dominate microbe. This is explained by there being more sugars in the "heavy beer", which gave the yeast the advantage. Acetic acid bacteria were still not detected during secondary fermentation <ref name="Martens"></ref>. During secondary fermentation, ''L. delbruekii ssp delbruekii'' strains dominated over the other lactic acid bacteria found. Additional strains of ''L. plantarum'' and ''L. coryneformis'', and an additional strain of ''L. brevis'' was found in the "heavy beer". Other than the dominating ''L. delbruekii ssp delbruekii'' strains, only a few strains of ''L. brevis'' were found in the "light beer" <ref name="Martens"></ref>. The "light beer" appeared to have a a smaller diversity of microbes.  ====Tertiary Fermentation====The same study by Martens et al. looked at two casks during the third fermentation, where the "heavy beer" beer was transferred from the secondary fermentation vessel for long term aging. At the beginning of the third phase of fermentation, ''Saccharomyces'' cell counts began to drop while the appearance of ''Brettanomyces'' began. After 10 weeks in the casks, ''Lactobacilli'' greatly decreased, giving rise to strains of ''Pediococcus parvulus''. After 12 weeks for Cask A and 18 weeks for Cask B, the beer no longer contained ''Saccharomyces'', and ''Brettanomyces'' dominated. Specifically, ''B. lambicus'' (now classified as a strain of ''B. bruxellensis'') and ''B. bruxellensis'' were the dominate species, but much smaller counts of ''B. intermedius'' (now classified as ''B. anomala'') and ''B. custersianus'' were also found. The ''Brettanomyces'' species continued to be the primary microorganisms for 36 weeks in Cask A and 50 weeks in Cask B. After this time period, ''P. parvulus'' began to dominate. Acetic acid bacteria also began to make an appearance in the casks, being detected at 27 weeks in Cask A and 40 weeks in Cask B. The exact numbers of the acetic acid bacteria were not reported by this study since they mostly grow on the surface of the beer inside the cask, and possibly on the walls of the cask where diffused oxygen could reach them more easily, and samples were not taken from these sections of the casks. The difference in the time periods for the microbial populations was determined to be affected by the casks themselves, which differed in age, size, and possibly different microbe colonization inside them before they were filled <ref name="Martens"></ref>.  ====Comparison to Lambic====Lambic is a similar beer produced in Belgium, but is fermented using [[Mixed Spontaneous Fermentation|mixed culture spontaneous fermentation]]. Enterbacteria were not found in the brewery that Martens et al. studied since spontaneous fermentation was not used. However, after the enterobacteria and primary ''Saccharomyces'' fermentation phases of lambic brewing are complete, the microbial populations of lambic and Flanders red/brown beers are similar during their aging processes. Both beers display a dominance by ''Brettanomyces'' and ''Pediococcus'' during the aging phases. Flanders red ales differ by having a large portion of the acid production performed by ''Lactobacilli'', where as in lambic the acid production is performed by ''Pediodoccus damnosus''. Flanders red ales are also characterized by having ''P. parvulus'' instead of ''P. damnosus'', however Martens et al. noted that the two species have no clear difference as far as their effects on fermentation go. Martens also noted that ''Brettanomyces'' began to disappear from old English Porter when the beer moved from wooden casks to metallic ones. It is thought then that the wooden casks are vital to ''Brettanomyces'' in Flanders Red Ale brewing, perhaps due to the presence of ''Pediococci'', with which ''Brettanomyces'' "cooperates" to ferment dextrins in the beer during the aging phase <ref name="Martens"></ref>.