Changes

'''Lactobacillus''' (often referred to by brewers as "Lacto") is a genus of Gram-positive, rod-shaped lactic acid bacteria (LAB) which produces acidity and sour flavors in the form of lactic acid and [[Lactobacillus#Sugar_Utilization_and_Secondary_Metabolites|secondary metabolites]] found in lambics, Berliner Weiss, sour brown ales, and gueuze. All ''Lactobacillus'' species are facultative anaerobes, which means they grow anaerobically but can also grow in the presence of oxygen and use oxygen to some degree <ref name="todar_lactics4"></ref>. There are more than 100 species, many of which are found in the human gastrointestinal track <ref name="todar_lactics4">[http://textbookofbacteriology.net/lactics_4.html ''Lactic Acid Bacteria''. Todar's Online Texbook of Bacteriology. Kenneth Todar, PhD. Pg. 4. Retrieved 07/28/2015.]</ref><ref name="Todar_nutgro4">[http://textbookofbacteriology.net/nutgro_4.html ''Nutrition and Growth of Bacteria''. Todar's Online Texbook of Bacteriology. Kenneth Todar, PhD. Retrieved 07/28/2015.]</ref>. In addition to beer, some species of ''Lactobacillus'' are also used to ferment yogurt, cheese, sauerkraut, pickles, wine, cider, kimchi, cocoa, and kefir <ref>[https://en.wikipedia.org/wiki/Lactobacillus ''Lactobacillus''. Wikipedia. Retrieved 07/28/2015.]</ref>. ''Lactobacillus'' can form a [[pellicle]] (need reference). See ''[[Pediococcus]]'', ''[[Brettanomyces]]'', ''[[Saccharomyces]]'', and [[Mixed Cultures]] charts for other commercially available cultures. See the [[Sour Worting]] and [[Mixed Fermentation]] pages for brewing techniques with ''Lactobacillus''. See the [[Alternative Bacteria Sources]] section for culturing ''Lactobacillus'' from grains, yogurt, probiotics, and other sources.

| Lallemand || WildBrew Sour Pitch || L. plantarum <ref>[https://www.facebook.com/Lallemandyeasts/photos/a.941604692537326.1073741829.939455986085530/1656901501007638/?type=3&comment_id=1657229347641520&reply_comment_id=1657231934307928&comment_tracking=%7B%22tn%22%3A%22R5%22%7D Post on the Lallemand Facebook page. 09/22/2017. Retrieved 09/22/2017.]</ref> || Facultatively homofermentative || || See [https://www.facebook.com/groups/MilkTheFunk/permalink/1790290834332456/ this information from Scott Lucas on MTF]. This culture comes in a dry (desiccated) format. Although the [http://www.lallemandbrewing.com/product-details/wildbrew-sour-pitch manufacturer's website] claims this strain is tolerant of 8 IBU, we recommend that brewers treat this strain like any other strain of ''L. plantarum'' and do not expose it to any hops until the desired acidity has been produced (for example, see [[Wort Souring]]). Recommended temperature: 86-104°F.

| [[The Yeast Bay]] || TYB282 || L. brevis || Heterofermentative || || TYB282 is a single strain of Lactobacillus brevis isolated out of an unintentionally soured golden ale produced by a Mexican craft brewery.This strain produces a nice, clean lactic acidity (down to ~pH 3.16-3.18) in unhopped wort within 36 hours at a temperature of ~72-77 F. The higher the temperature (up to 90 F is what we've tested), the faster the acid production. This is a great strain for kettle souring, as it grows rather quickly and produces acidity fast with no detectable off flavors. We are about to begin some trials in hopped wort to test out the acidification in the presence of hop compounds, though we presume there is some level of adaptation to hop compounds given the environment in the beer from which we cultured the strain. Temperature: 70-90 ºF. |-| [[White Labs]] || WLP677 || L. delbrueckii (potentially misidentified) || Heterofermentative <ref name="mtf_wiki_shaner">[http://www.milkthefunk.com/wiki/100%25_Lactobacillus_Fermentation Milk The Funk Wiki. 100% Lactobacillus Fermentation Test by Lance Shaner.]</ref><ref name="tmf_cultures">[http://www.themadfermentationist.com/p/commercial-cultures.html ''Commercial Brettanomyces, Lactobacillus, and Pediococcus Descriptions''. The Mad Fermentationist Blog. Michael Tonsmeire. Retrieved 3/4/2015.]</ref> || no stir plate, room temp ||Incubate at > 90°F and < 117°F for 5-7 days for greater lactic acid production. Cell count: 50-80 million cells/mL (1.75-2.8 billion cells in a 35 mL homebrew vial) <ref name="WL_cellcounts">Private correspondence with White Labs Customer Service and Dan Pixley. 10/29/2015.</ref>. Not a good strain for kettle souring, but can produce a "soft" acidity over a longer period of time <ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/1212455192116026/?comment_id=1212475888780623&reply_comment_id=1212476575447221&comment_tracking=%7B%22tn%22%3A%22R3%22%7D Conversation with Andrew Addkison on MTF. 01/12/2016.]</ref>. White Labs claims that it is tolerant to up to 20 IBU, although growth starts to become inhibited at 15 IBU <ref name="WL_datasheet" /><ref>[http://www.themadfermentationist.com/p/commercial-cultures.html "Commercial Brettanomyces, Lactobacillus, and Pediococcus Descriptions; Commercial Yeast Laboratories." The Mad Fermentationist blog. Michael Tonsmeire. Retrieved 12/12/2016.]</ref>. Generally heat tolerant, but sours faster between 100-110°F <ref name="WL_datasheet">[http://www.whitelabs.com/sites/default/files/R%26D%20Wild%20Yeast%20and%20Bacteria%20Experiments_2.pdf "R&D Wild Yeast and Bacteria Experiments". White Labs data sheet. Retrieved 05/16/2017.]</ref>

Although ''Lactobacillus'' are tolerant of oxygen and oxygen usually does not negatively affect their growth (except in the case of ''L. plantarum'', which has been shown to produce small amounts of acetic acid when exposed to oxygen and glucose is not present <ref name="Quatravaux_plantarum">[http://onlinelibrary.wiley.com/doi/10.1111/j.1365-2672.2006.02955.x/full Examination of Lactobacillus plantarum lactate metabolism side effects in relation to the modulation of aeration parameters. S. Quatravaux, F. Remize, E. Bryckaert, D. Colavizza, J. Guzzo. 2006]</ref><ref name="microbewiki_plantarum"></ref>), it is also generally not needed (an exception to this may be ''L. brevis'', which has been shown to increase growth rates in the presence of oxygen <ref name="brevis_aeration"></ref>). Therefore, it is generally best practice to prevent aerating the starter with an airlock for ''Lactobacillus'' starters. If exposure to air occurs, and the starter does not smell like it has been contaminated by the exposure, then the starter can still be used.

The recipe for this starter wort is: '''1.040 SG (10°P) Dried Malt Extract wort with 10% apple juice + 20 grams of chalk (CaCO3) per liter + yeast nutrients'''. Regarding the use of chalk, it is the preferred buffer because it does not react with CO2 (unlike baking soda), so it won't be consumed by exposure to air due to CO2 production by the Lacto. It also has a pKa (maximum buffering capacity) of around 4.6, which is ideal for ''Lactobacillus'' growth. The fact that it easily precipitates out also makes it ideal to use as a buffer <ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/1180630378631841/?comment_id=1181674265194119&reply_comment_id=1181743348520544&comment_tracking=%7B%22tn%22%3A%22R%22%7D Conversation with Bryan of Sui Generis Blog regarding the use of chalk as a buffer in Lacto starters. 11/20/2015.]</ref>. Jeff Mello from [[Bootleg Biology]] and , Nick Impellitteri from [[The Yeast Bay]], and Bryan from [https://suigenerisbrewing.blogspot.com/ Sui Generis blog] suggest that using the smaller amount of 1.5-2 grams of CaCO3 per liter is preferable because that amount is easier to precipitate out of the starter and avoid pitching into the beer (the growth differences from using less chalk has not been tested though) <ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/1369904163037794/?comment_id=1370329352995275&reply_comment_id=1372184639476413&comment_tracking=%7B%22tn%22%3A%22R%22%7D Conversation with Jeff Mello on MTF regarding using less chalk in LAB starters. 08/10/2016.]</ref><ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/1619935741367967/?comment_id=1619986154696259&reply_comment_id=1619991214695753&comment_tracking=%7B%22tn%22%3A%22R9%22%7D Impellitteri, Nick. Milk The Funk Facebook group. 03/19/2017.]</ref><ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/1457469187614624/?comment_id=1457541770940699&reply_comment_id=1457620514266158&comment_tracking=%7B%22tn%22%3A%22R%22%7D Bryan from Sui Generis. MTF Thread on using 1.5g/L of chalk for Lactobacillus starters. 11/02/2016.]</ref>. To create a 1 liter starter for 20 liters of wort, follow these directions:

<blockquote>"I typically grow it by itself anaerobically in [http://www.neogen.com/Acumedia/pdf/ProdInfo/7406_PI.pdf MRS media]. Seems to work very well and results in good growth. I've personally had the best success with MRS media and in an anaerobic environment, though I know some ''Lactobacillus'' strains grow aerobically just fine. The problem with growing lactic acid bacteria is the acid they produce will eventually inhibit their own growth. MRS contains a buffer to help combat the drop in pH as a result of LAB metabolism, which keeps the pH around 6-6.5 (I think) for optimal growth. I usually grow them at 35 C, but sometimes incubator space is at a premium (like right now) and I just [use a stir plate with an airlock]" <ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/1031115430250004/?comment_id=1031228363572044&offset=0&total_comments=24 Conversation with Nick Impellitteri on Milk The Funk Facebook group. 3/5/2015.]</ref>. - Nick Impellitteri from [[The Yeast Bay]] on general Lactobacillus cell growth</blockquote>

All [https://en.wikipedia.org/wiki/Prokaryote prokaryotes], which includes all bacteria, are categorized based on the levels of oxygen in their environment in which they can grow and how they utilize oxygen, if at all <ref name="Todar_nutgro4"></ref>. ''Lactobacillus'' species are usually considered to be "facultative anaerobes" (or "facultative aerobes") <ref name="todar_lactics4"></ref>, however they are a special case. Facultative anaerobes usually make energy from oxygen if it is present via the [https://en.wikipedia.org/wiki/Oxidative_phosphorylation oxidative phosphorylation pathway], but otherwise engage in anaerobic fermentation <ref>[http://www.ncbi.nlm.nih.gov/books/NBK21208/ Biochemistry. 5th edition. Berg JM, Tymoczko JL, Stryer L. 2002. Chapter 18.]</ref><ref>[http://inst.bact.wisc.edu/inst/index.php?module=book&type=user&func=displayarticle&aid=111 Virtual Microbiology Textbook. Department of Bacteriology, University of Wisconsin-Madison. Retrieved 12/02/2015.]</ref>. ''Lactobacillus'' species can utilize oxygen, but not through the oxidative phosphorylation pathway. They use an alternative pathway instead. This pathway uses flavine-containing oxidases and peroxidases to carry out the oxidation of NADH2 using O2 <ref name="bergey">Bergey's Manual of Systematic Bacteriology, 2nd edition. pg 471</ref><ref>Correspondence with Bryan of Sui Generis Blog from Dan Pixley. 12/01/2015.</ref>. Lactobacilli, therefore, are unique in that they blur the line between facultative anaerobes and another class of prokaryotes known as "aerotolerant anaerobes". Aerotolerant anaerobes do not use oxygen to generate energy, but can grow in the presence of oxygen.

There are, however, a few exceptions to this in the scientific literature. For example, ''L. plantarum'', which is a facultatively heterofermentative species, is homolactic without the presence of oxygen and produces only lactic acid. After food sources have been exhausted and in the presence of oxygen, however, ''L. plantarum'' switches to heterolactic fermentation, and further converts lactic acid into acetic acid. In a lab setting the conversion of lactic acid to acetic acid only happened when glucose was no longer available (this may not be the case in wort where other limiting factors such as low pH can prevent ''L. plantarum'' from continuing their metabolic processes before all glucose is consumed, which is the case in kettle souring), and only during the stationary phase (after growth stopped). During the conversion of lactic acid to acetic acid, ''L. plantarum'' also produces hydrogen peroxide (H²0²), which is toxic to microorganisms and is thought to be a protection mechanism for ''L. plantarum'' <ref>[http://www.ncbi.nlm.nih.gov/pubmed/6480562 Physiological role of pyruvate oxidase in the aerobic metabolism of Lactobacillus plantarum. Sedewitz B, Schleifer KH, Götz F.1984.]</ref><ref name="Quatravaux_plantarum"></ref><ref name="microbewiki_plantarum">[https://microbewiki.kenyon.edu/index.php/Lactobacillus_plantarum_and_its_biological_implications Lactobacillus plantarum and its biological implications. Microbe Wiki. Retrieved 6/7/2015.]</ref><ref name="shaner_plantarum"></ref>. This same process has also been observed in one strain of ''L. brevis'' and under similar conditions of depleted nutrients and oxygen <ref>[http://aem.asm.org/content/early/2017/08/21/AEM.01659-17.abstract The oxygen-inducible conversion of lactate to acetate in heterofermentative Lactobacillus brevis ATCC367. Tingting Guo, Li Zhang, Yongping Xin, ZhenShang Xu, Huiying He, and Jian Kong. 2017.]</ref>. Omega Yeast Labs reports that no noticeable acetic acid is produced if the oxygen is not purged with their OYL-605 ''Lactobacillus'' blend which contains ''L. plantarum'', and brewers should not aerate wort during sour but purging O2 is not required <ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/1222363364458542/?comment_id=1222371257791086&reply_comment_id=1225212894173589&comment_tracking=%7B%22tn%22%3A%22R%22%7D MTF conversation with Adi Hastings from Omega Yeast Labs on the effects of oxygen presence in OYL-605. 02/02/2016.]</ref>. This indicates that acetic acid and hydrogen peroxide production in typical brewing scenarios where little or no oxygen is present is probably not a concern. ''L. brevis'' has been shown to increase growth rates in the presence of oxygen <ref name="brevis_aeration"></ref>. Thomas Hübbe's master thesis showed that under lab growth media "VLB-S7-S", "NBB®-A", and MYPG + cyclohexmide, both strains of ''Lactobacillus'' tested (''L. brevis'' and ''L. parabrevis'') did not show growth in an aerobic chamber, but did grow in an anaerobic chamber , although this might be an effect of the growth media and not representative of brewing conditions <ref name="Hubbe">[https://www.facebook.com/groups/MilkTheFunk/1407620505932826/ Effect of mixed cultures on microbiological development in Berliner Weisse (master thesis). Thomas Hübbe. 2016.]</ref>.

[http://www.sciencedirect.com/science/article/pii/S0308814617302911#t0005 Dongmo et al. (2017)] found 56 volatile flavor compounds, including various esters, alcohols, ketones, aldehydes, acids, ethers compounds, sulfur compounds, heterocyclic compounds, phenols(including guaiacol and 4-vinylguaiacol), terpenes, lactones, and several unidentified compounds. Key compounds produced by ''Lactobacillus'' include acetaldehyde (thought to be a major flavor contributor to kettle soured beers <ref name="Peyer_2017" />), β-Damascenone, furaneol, phenylacetic acid, 2-phenylethanol, 4-vinylguaiacol, sotolon, methional, vanillin, acetic acid, nor-furaneol, guaiacol and ethyl 2-methylbutanoate. Acetaldehyde was the most impactful aroma compound found followed by propan-1-ol and γ-dodecalactone. Acetaldehyde was generally produced in much higher amounts (~23-64 µg/L) by the select strains of ''L. plantarum'', while ''L. amylolyticus'' and ''L. brevis'' produced only 1.5-3 µg/L. In fact, the levels of all of these compounds differed significantly based on the species and strain. The selected strains of ''L. brevis'' were associated as having worse aromas that were dominated by methional (cooked potatoes), acetic acid (vinegar), and nor-furaneol (caramel-like). The ''L. plantarum'' strains selected were identified as producing more positive aromas from compounds such as β-damascenone (apple/fruit juice), furaneol (strawberry), 2-phenylethanol (rose/caramel) and ethyl 2-methylbutanoate (citrus) Small but significant amounts of linalool and geraniol were also found, which are normally terpenes found in [[Hops|hops]]. Vanillan is formed from ferulic acid by some ''Lactobacillus'' species as well as ''Oenococcus oeni'' <ref name="Dongmo">[http://www.sciencedirect.com/science/article/pii/S0308814617302911 Key volatile aroma compounds of lactic acid fermented malt based beverages – impact of lactic acid bacteria strains. Sorelle Nsogning Dongmo, Bertram Sacher, Hubert Kollmannsberger, Thomas Becker. 2017. doi:http://dx.doi.org/10.1016/j.foodchem.2017.02.091.]</ref>.